Scandem Book of Abstracts 25 June 2018

SCANDEM 2018 Book of Abstracts

69 Annual th

Meeting of the Nordic Microscopy Society June 25th–28th, 2018 Technical University of Denmark Copenhagen, Denmark

Scandem2018.com

Nordic Microscopy Society Scandem2018.com

SCANDEM 2018 26-28 June 2018 Technical University of Denmark

BOOK OF ABSTRACTS Organizer Jakob Birkedal Wagner

Linking Nordic Microscopists to promote new technology, techniques and applications

4

TABLE OF CONTENTS Abstracts Plenary Session ……..…………………………………………….………….…...... 7-9 Keynote Session ……..…………………………………………….………….…..10-16 Materials Science and Energy Materials ………..……………….………….…..17-46 In Situ Nanoscale Microscopy of Processes ………..….……….………….…. 47-72 Advances in electron spectroscopy: techniques, instrumentation and applications .................................................................................................... 73-90 Image Data & Analysis .................................………..….……….………….…. 91-106 Imaging multicellular systems, Live imaging of single cells, Correlative Light and Electron Microscopy (CLEM) ....................................................... 107-120 Author Index .................................………..….…..................…….…………...121-126

5 ORGANIZING COMMITTEE Conference Chair:

Exhibition:

Jakob Birkedal Wagner

Andrew Burrows

Members:

Berit Wenzell

Nina Høgh-Bach Kirsten Noelle Bruus

Michael Andersson (Bruker Nano Analytics)

Mette Noer

Torben Jensen (Fischione)

SCIENTIFIC PROGRAM COMMITTEE Thomas Willum Hansen (DTU)

Sebastian Horch (DTU)

Jacob R. Bowen (DTU)

Anders Bjorholm Dahl (DTU)

Carsten Gundlach (DTU)

Kristian Mølhave (DTU)

Michael Lisby (KU)

Rodolphe Marie (DTU)

Morten Schallburg Nielsen (AAU)

Shima Kadkhodazadeh (DTU)

STUDENT Volunteers Mamadou Mandie Kone (DTU)

Anton Bay Andersen (DTU)

Mohammad Ahmed (DTU)

Asger Barkholt Moss (DTU)

Thomas L. Smitshuysen (DTU)

6 SCANDEM Board of Directors The Executive Board President Kesara Anamthawat-Jónsson Plant Genetics Research Group, Faculty of Life and Environmental Sciences, School of Engineering and Natural Sciences, University of Iceland

Vice President and Secretary General Varpu Marjomäki Department of Biological and Environmental Science, University of Jyväskylä

Treasurer Salla Marttila Department of Plant Protection Biology, Swedish University for Agricultural Sciences (SLU)

Information Officer Lassi Paavolainen

Company Representative Michael Andersson

Institute for Molecular Medicine Finland

Bruker Nano Analytics, Sweden

Representatives from Denmark Søren Fæster Nielsen

Klaus Qvortrup

Materials Research Department, Risø DTU National Laboratory for Sustainable Energy

Department of Biomedical Sciences/CFIM, University of Copenhagen

Representatives from Finland Eija Jokitalo

Minnamari Vippola

Institute of Biotechnology, Electron Microscopy Unit, University of Helsinki

Materials Characterization Group, Department of Materials Science, Tampere University of Technology

Representative from Iceland Kristjan Leosson Innovation Centre Iceland

Representatives from Norway Randi Holmestad

Ingunn Thorseth

Department of Physics, Norwegian University of Science and Technology

Department of Earth Science and Center for Geobiology, University of Bergen

Representatives from Sweden Per Persson

Oleg Shupliakov

Thin Film Physics Division, Department of Physics, Chemistry and Biology, Linköping University

Neuronal Membrane Trafficking, Center of Excellence in Developmental Biology, Karolinska Institutet

Session: Plenary Catalysis: A Key Materials for Converting Power into Chemicals and Fuels.

Ib Chorkendorff SurfCat, Department of Physics, Technical University of Denmark (DTU) Fysikvej, Building 312, DK-2800 Kongens Lyngby, Denmark.

E-mail: ibchork@fysik.dtu.dk Keywords: heterogeneous catalysis, solar fuels, electrolysis, nanoparticles In this presentation, I will give an overview of the need to produce new and effective catalysts for energy conversion. The approach and methods shall be discussed in the light of know limitations defining the way forward [1]. We shall for example demonstrate how mass-selected nanoparticles, can be used to elucidate the activity for processes related to electrolysis and the reversible process in fuel cells. Here the Oxygen reactions are the cause of limited activity. It shall be shown how an entirely new classes of electro-catalysts have been identified by alloying Pt with early transition metals or the lanthanides [2]. Here mass-selected nanoparticles have been essential for demonstrating that high area materials can display high activity and allowing for gaining insight into the fundamental cause of the strong size dependent activity [3]. We shall demonstrate how similar approach can be used for studying size dependence and efficiency for catalysts related to water electrolysis [4], showing how isotope labeling can help determining the activity of bulk versus surface.

Having hydrogen, it is possible to synthesize both fuels and chemicals by hydrogenating CO2. We shall demonstrate how it is possible to identify the active site of the methanol catalysts and based on such insight make prediction for the optimal catalysts [5]. It shall be demonstrated how a similar route for synthesizing small organic molecules can be done electrochemically.

This approach has been used to investigate the dynamical influence of surface oxygen on electrochemical CO hydrogenation into methane/ethene on mass-selected Copper nanoparticles [6].

References: [1] Z. W Seh, J. Kibsgaard, C. F. Dickens, I. Chorkendorff, J. K. Nørskov, T. F. Jaramillo, Science 355 (2017) [2] M. Escudero-Escribano, … I. E.L. Stephens, I. Chorkendorff, Science 352 73 (2016).

[3] P. Hernandez-Fernandez, , …, I. Chorkendorff, Nature Chemistry 6 732 (2014).

[4] C. Roy, …J. Kibsgaard, I. E. L. Stephens, and I. Chorkendorff, Submitted (2018).

[5] S. Kuld, M. Thorhauge, H. Falsig, C. Elkjær, S. Helveg, I. Chorkendorff, J. Sehested, Science 352 969-974 (2016).

[6] S. B. Scott, …. P. C. K. Vesborg, Jan Rossmeisl, and I. Chorkendorff, Submitted (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark P-1

1

Session: Plenary Vizualising Bacteria and their Behaviour Lone Gram Department of Biotechnology and Biomedicine, Technical University of Denmark, DK-2800 Kgs. Lyngby E-mail: gram@bio.dtu.dk Keywords: anti-bacterial compounds, beneficial bacteria, novel bacterial species Microorganisms are of immense importance for mankind and influence all areas of life. They are responsible for turn-over of nutrients in the biogeochemical cycles, some cause infectious diseases, many contribute to the disease-defense of eukaryotes and they are invaluable sources of drugs and bioprocessing compounds as well as serving as cell-factories in the biomanufacturing industry. They are tiny; in the micrometer range and invisible to the naked eye.

The invention and introduction of devices and instruments for enlargement; ranging from van Leeuwenhoek’s first microscopes in the 16th hundreds to current day high resolution microscopes based on electron beams or atomic force has truly revolutionized microbiology.

This talk will present examples of work aimed at controlling or using microorganisms and present how visualization of the microorganisms can facilitate understanding of the systems. (i) Identification: Microorganisms are as classified in taxonomic systems with the species as the core unit. Identify and classifying microorganisms correctly is important in both clinical microbiology and biotechnology. With the development of methods allowing rapid genetic and metabolic characterization of microorganisms, we have come to realize that the diversity is unsurpassed and that 1,000s of novel species are “out there�. As part of identifying and naming a novel species, high resolution microscopy pictures are very often included (Fig 1, [1]). (ii) Novel antimicrobials: Treatment of infectious diseases was improved dramatically with the discovery of penicillin and other antibiotics, however, the rapid development and spread of antibiotic resistance has led to an urgent need for novel antimicrobials. Determining the mechanism of action (MoA) is required to get approval of novel drugs and visualizing (SEM, AFM a.o.) of the actual action of novel compounds facilitates this process (Fig 2, [2]).

Preventing transmission of pathogenic agents is key in limiting spread of infections. Bacteria prefer to attach to surfaces (abiotic and biotic) and in, for instance, hospitals, inert surfaces such as door handles, bed railings, water knobs can act as vectors of transfer. Surfaces with antibacterial effects is an important intervention strategy, however, the antibacterial principle should not select for resistant organisms. One example is a novel Cu-Ag alloy developed by Elplatek A/S. Fluorescence microscopy of living and dead cells enables an in situ assessment of killing kinetics and is used in subsequent documentation (Fig 2, [3]). (iii) Beneficial bacteria: The transmission of pathogenic bacteria must also be controlled in food production, and, as in the clinical sector, preferably with novel antibiotic independent strategies. Microscopy can assist in identifying sites where pathogens colonize and thus where interventions should focus. In fish production, the live feed (algae, copepods) can serve as transmission vectors for pathogens (Fig. 3 [4,5]) and their colonization and growth can be prevented by introduction of probiotic bacteria that can inhibit the pathogen with no adverse effect on the host (the live feed) (Fig. 3 [4,5]).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark P-2

Session: Plenary

Figure 1: A novel species: Vibrio galathea as seen in high vacuum SEM (left) and environmental EM (100% humidity) (middle) [1].

Figure 2: Mechanism of action of antibacterials. Antimicrobial peptides causing membrane damage and leakage from E. coli (left: un-treated cells; middle: treated cells). CuAg-surface causing contact killing of S. aureus (right: red: dead cells; green: live cells) [2,3]

Figure 3: Light- and fluorescence microscopy of colonization of live fish feed (copepods) with a GFP labelled fish pathogen [4] and of algae with beneficial, probiotic bacteria [5].

[1] S. Giubergia, H. Machado, R.V. Mateiu and L. Gram. Int J System Evol Microbiol 66, 347–352 (2016) [2] R.V. Mateiu, L. Citterio and L. Gram. SEM of Escherichia coli treated with antimicrobial peptides. Un-published data (2016) [3] N. Ciacotich and K.N. Kragh. Confocal scanning microscopy of Staphylococcus aureus on a Cu-Ag-surface. Unpublished data. (2018) [4] B.B. Rasmussen, K.E. Erner, L. Gram and M. Bentzon-Tilia. Microb. Biotechnol. In revision (2018) [5] P. D’Alvise, S. Lillebo, M.J.P. Garcia, H.I. Wergeland, K. F. Nielsen, Ø. Bergh and L. Gram. PloS One 7:e43996 (2012)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark P-2

11

Session: Keynote Correlative Imaging: From Cells to Stars Lucy M. Collinson*1 1

Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK. *E-mail: lucy.collinson@crick.ac.uk

Keywords: correlative light and electron microscopy, cell biology, astrophysics, big data, citizen science.

Correlative light and electron microscopy (CLEM) combines the benefits of fluorescence and electron imaging, revealing protein localisation against the backdrop of cellular architecture.

The correlative imaging field is expanding rapidly, and encompasses workflows that link many different imaging modalities, to answer scientific questions in the biological and physical sciences. We link fluorescence microscopes (widefield, confocal, super-resolution and lightsheet) with electron microscopes (scanning, transmission, serial block face and focused ion beam) [1] and X-ray microscopes (microCT and soft X-ray) to analyse a range of biological samples, from single cells to whole model organisms.

Our technology development work has focused on improving the speed, accuracy and accessibility of CLEM. During this development work, it became clear that the technical challenges associated with correlative imaging are exaggerated when working in 3D [2]. To increase protein localisation precision, we developed an ‘In-Resin Fluorescence’ (IRF) protocol that preserves the activity of GFP and related fluorophores in resin-embedded cells and tissues.

The sample preparation is relatively fast, and also introduces electron contrast so that cell structure can be visualised in the electron microscope. Once the resin blocks have been cut into ultrathin sections, out-of-plane fluorescence is removed resulting in physical ‘super-resolution’ light microscopy in the axial direction, which increases the accuracy of the LM-EM overlays.

Localisation precision is further increased by imaging the IRF sections in vacuo in the next generation of commercial integrated light and electron microscopes (ILEM). We were able to further improve accuracy by developing integrated super-resolution light and electron microscopy, using the remarkable blinking properties of GFP and YFP in-resin in vacuo [3].

With the advent of dual contrast samples comes the potential to locate and track fluorescent cells during sample preparation and automated 3D EM image acquisition. We designed and built two new locator tools – a fluorescence microscope designed to integrate with an ultramicrotome to locate cells during trimming and sectioning (the ultraLM), and an even smaller version that fits into the extremely tight space of the SBF SEM vacuum chamber for on-the-fly tracking of fluorescent cells during long automated imaging runs (the miniLM) [4].

As electron microscopes become more automated, data outputs are increasing astronomically, and so the bottleneck in our work is shifting from data acquisition to data analysis. I will describe the ideas and workflows we are developing to deal with big data, and describe our Citizen Science collaboration with the Zooniverse platform, which is helping us to develop automated detection and segmentation of cell structures in EM images.

[1] C. Peddie & L.M.Collinson, Micron 61, 9-19 (2014).

[2] M. Russell et al., JCS 130, 278-91 (2016).

[3] C. Peddie et al., J Struct Biol 199, 120-31 (2017).

[4] E. Brama et al., Wellcome Open Research 1, 26 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark K-1

Session: Keynote Visualizing the Atomic-scale Structure and Dynamics on Catalyst Materials using Scanning Probe Microscopy Jeppe V. Lauritsen.

1

Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Denmark.

E-mail: jvang@inano.au.dk Keywords: scanning probe microscopy, oxide surfaces, diffusion, catalysis.

The development of new materials for catalysis is seen as a crucial progress for securing energy resources and for better protection of the environment. Obtaining detailed control of materials on the nanoscale is of essence in catalyst development, but often a lack of insight into the fundamental physical and chemical processes occurring on catalytically active surfaces hampers the progress. We pursue the goal of understanding catalytic processes on surfaces by focusing on what happens on the atomic level. Scanning Probe Microscopy techniques (SPMs) are particularly strong techniques in this regard, since they allow us to visualize the atomic structure of surfaces and sometimes directly see the outcome of catalytic reactions. In my talk, I give examples on how we successfully use the scanning tunneling microscope (STM) in interplay with other surface science techniques to investigate industrially used catalysts for e.g.

NOx pollution abatement and new earth-abundant catalyst materials for H2 production [1-3].

For both these catalysts, STM has successfully revealed the catalytic importance of point defects, which are otherwise difficult to study by other means than microscopy. Furthermore, time-lapsed STM movies are used to reveal the atomistic mechanisms involved in surface diffusion and reactions. Applying catalytic conditions at elevated pressure can lead to important changes in the surface structure of materials, and I will also briefly outline how such challenges can be met by new scanning probe microscopy instrumentation capable of imaging and characterizing surfaces while the catalyst is under ‘working conditions’.

Figure 1: A cobalt oxide island imaged with atom-resolved scanning tunneling microscopy while reacting with gas phase H2O..

[1] [2] [3]

J. Fester, M. García-Melchor, A. S. Walton, et al., Nat. Commun. 8, 14169 (2017).

S. Koust, L. Arnarson, P. G. Moses, et al., Phys. Chem. Chem. Phys 19, 9424 (2017).

S. Koust, B. N. Reinecke, K. C. Adamsen, et al., Journal of Catalysis 360, 118 (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark K-2

Session: Keynote X-ray Microtomography – A Versatile Tool F Ahmed1,2 Exponent International, London UK 2 Imaging and Analysis Centre, The Natural History Museum, London UK. 1

.

E-mail: fahmed@exponent.com Keywords: XMT, Imaging Techniques, Applications.

X-ray microtomography (XMT) has evolved exponentially as an imaging technique. XMT is used in all aspects of academia and industry, including research studies, product inspections, manufacturing processes and failure analysis. Over the last decade, XMT has been used to address some of the most complex problems facing the natural world. Its role in creating digital libraries in museums, designs for additive manufacturing and visual appeal in engaging the public has led to global awareness.

Cultural shifts in sharing data and open access repositories allow communities around the world to access unprecedented information, leading to ground-breaking discoveries. This talk will focus on specific case studies, where collaboration on a global scale has aided researchers and members of industry to solve real-world problems. .

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark K-3

Session: Keynote Balancing Spatial, Energy and Momentum Resolutions in Electron Energy Loss Spectroscopy Quentin M. Ramasse*1,2, Fredrik S. Hage1, Rebecca J. Nicholls3, Jonathan R. Yates3, Demie M. Kepaptsoglou1, Trevor P. Hardcastle2, Morten N. Gjerding4 and Kristian S. Thygesen4 1

SuperSTEM, SciTech Daresbury Campus, Keckwick Lane, Daresbury WA4 4AD, U.K. School of Chemical and Process Engineering, University of Leeds, Leeds LS2 9JT, U.K. 3 Department of Materials, University of Oxford, Parks Road, Oxford OX1 3PH, U.K. 4 CAMD and Center for Nanostructured Graphene (CNG), Denmark Technical University, Fysikvej 1, building 307, 2800 Kgs. Lyngby, Denmark *E-mail: qmramasse@superstem.org 2

Keywords: scanning transmission electron microscopy, electron energy loss spectroscopy, carbon nanomaterials, momentum resolution.

The properties of materials are increasingly controlled and tuned through defects engineering taking place quite literally at the atomic level, where one of the most powerful means of characterization arguably lies within a combination of low voltage scanning transmission electron microscopy, energy loss spectroscopy (STEM-EELS) and ab initio calculations.

This approach can for instance be used to unambiguously confirm the p and n character induced in graphene through the inclusion of single atom B or N substitutional dopants, achieved by low energy ion implantation technology akin to that routinely exploited in the semi-conductor industry [1]. The inclusion of single B and N dopants in the lattice is also shown to induce a dampening or enhancement of the characteristic interband π plasmon of graphene through a high-resolution EELS study carried out on a new generation of STEM, a monochromated Nion UltraSTEM100 MC capable of an energy resolution down to 14 meV [2]. A relative 16% decrease or 20% increase in the π plasmon quality factor is attributed to the presence of the B or N atom dopants (Fig. 1). This modification is shown to be relatively localized and can no longer be detected beyond approximately 1nm from the dopant [3].

Extensive ab initio calculations using the local density approximation (LDA) exchangecorrelation functional as implemented in the electronic structure code GPAW, confirm the trends observed experimentally. Importantly, to achieve meaningful results that can be unambiguously compared with experimental data, it was found necessary to include the integration over momentum space required by the converged probe experimental geometry.

In this context, the EEL signal momentum transfer dependence (i.e. dispersion) can provide information that is often obscured in spectra acquired using experimental geometries optimized for high spatial resolution. However, momentum-resolved measurements, in particular in the vibrational spectroscopy regime, have so far been limited to “bulk” techniques (e.g. inelastic x-ray and neutron scattering spectroscopies), the average surface response (e.g. reflection EELS) or small momentum transfers (e.g. optical techniques).

We therefore developed a highly flexible combined experimental and theoretical methodology for acquiring and interpreting EEL signal at sub nm spatial resolution across the first Brillouin zone (BZ) in the electron microscope. Carefully balancing the intrinsic trade-off between simultaneously achievable resolutions in real and momentum space, we map the vibrational response of two polymorphs of boron nitride (BN) along different directions of their respective BZs, using a ~1nm electron probe. Effectively, we acquire spectra akin to those of inelastic X-ray and neutron spectroscopies while probing a sample volume that is ~1010-1020 times smaller [4]. The experimental data show remarkable agreement with ab

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark K-4

Session: Keynote initio calculations; acoustic (A), optical (O) and anisotropic phonon mode contributions are clearly identifiable in the experimental spectra.

A similar approach is applied to the low loss response of individual single-walled carbon nanotubes (CNTs), which contains a variety of characteristic losses, such as van Hove singularities and other interband transitions, in addition to plasmons, all of which are of great importance to understand their electronic behaviour and the role of defects in their electronic structure. A momentum-resolved study of the dispersion of the π plasmon of individual CNTs here highlight a variable π plasmon confinement parallel to the tube axis, which can be attributed to the a varying concentration of topological defects (i.e. non-hexagonal rings incorporated in the tube wall) within the individual CNTs [5].

Figure 1. (a) Medium angle annular dark field (MAADF) integrated intensity line profile compared to the local change in π plasmon peak width (FWHM) for N-doped graphene. A smoothed FWHM line profile is superimposed; (b) corresponding MAADF image; (c) π plasmon peak FWHM maps (image width: 1.1nm). The regions from which the line profile was extracted is indicated on (c). The localisation of the observed π plasmon enhancement (N) is estimated to be ~1 nm, as indicated.

Figure 2. (A) High angle annular dark field (HAADF) image of a single wall CNT. (B) low loss spectra acquired from (1) and (2) in A. (C) Momentum-resolved spectra. Smoothed data (red) is superimposed on zero loss subtracted raw data (grey line) as a guide to the eye.

[1] DM Kepaptsoglou et al., ACS Nano 9, pp.11398-11407 (2015).

[2] QM Ramasse, Ultramicroscopy 180, 41-51 (2017).

[3] FS Hage et al., ACS Nano 12, 1837-1848 (2018).

[4] FS Hage et al., Submitted (2018).

[5] FS Hage et al., PRB 95, 195411 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark K-4

Session: keynote Versatile image processing with Python and scikit-image, application to ultrafast in situ microtomography Emmanuelle Gouillart*1, 2 1

Joint Unit CNRS/Saint-Gobain Surface of Glass and Interfaces, 39 quai Lucien Lefranc, 93303 Aubervilliers, France 1 scikit-image development team *E-mail: emmanuelle.gouillart@saint-gobain.com Keywords: image processing, Python, X-ray tomography

While different imaging modalities push further the limits of spatial resolution or chemical sensitivity, extracting scientific information from image processing remains an important challenge. The reasons are manifold: images acquired in difficult conditions suffer from several artifacts or low signal over noise ratio. Furthermore physicists, chemists or biologists often lack academic background in mathematics and programming.

The talk will focus on the image processing capabilities of scikit-image, an open-source image processing library for the Python language. Scikit-image offers a variety of image processing operations, both for 2D and 3D images, so that it is compatible with most imaging modalities. A few examples, among many others, are given in Fig. 1: denoising noisy images, segmentation of objects, measuring their properties, etc. In addition, the package has a strong focus on thorough documentation, with innovative tools such as galleries of examples or interactive notebooks. Students and beginners in image processing can find their way through the documentation and learn the building blocks of image processing by example. Finally, scikit-image is part of the vibrant Scientific Python ecosystem, which makes it easy to interface classical image processing with other tasks such as machine learning, 3D visualization, etc.

Figure 1: Typical image processing operations with scikit-image. Both basic and more advanced algorithms are available for most processing tasks.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark K-5

Session: keynote I will illustrate the use of these tools with in situ imaging studies of materials at high temperature, using ultrafast synchrotron microtomography. Such experiments typically require to process hundreds of gigabytes of 3-D volumes, which have a low signal to noise ratio due to the fast acquisition rate. It is challenging to keep an agile and exploratory workflow for such large data sizes: tools for parallelization and caching of intermediate results still make it possible. I will present typical image processing pipelines which are successful for object segmentation and characterization despite strong noise and artifacts.

[1] Van der Walt, Stefan, et al. "scikit-image: image processing in Python." PeerJ 2 (2014): e453.

[2] Gouillart, E., Nunez-Iglesias, J., & Walt, S. (2017). Analyzing microtomography data with Python and the scikit-image library. Advanced Structural and Chemical Imaging, 2(1), 18.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark K-5

Session: Materials Science and Energy Materials Scanning Transmission Electron Microscopy of Polar Nano-regions and Atomic Scale Chemical Composition Fluctuations Study in Paraelectric (Ba, Sr)TiO3 Ceramics Goran Drazic*1, Andreja Bencan2, Tadej Rojac2 and Dragan Damjanovic3 1

National Institute of Chemistry, Hajdrihova ulica 19, Ljubljana, Slovenia.

2 Jozef Stefan Institute, Jamova ulica 39, Ljubljana, Slovenia.

3 Swiss Federal Institute of Technology - EPFL, Route Cantonale, Lausanne, Switzerland. *E-mail: goran.drazic@ki.si Keywords: functional oxides, ferroelectrics, polar nano-regions, scanning transmission electron microscopy, quantitative HAADF. Ferroelectric (Ba,Sr)TiO3 (BST) based materials undergo at a Curie temperature (Tc) a phase transition from a non-centrosymmetric polar ferroelectric phase to a paraelectric phase.

This phase is centrosymmetric (cubic) and the polarization is lost. However, recently it have been shown that cubic BaTiO3 phase exhibits breaking of nominal centric symmetry and exhibits polarization which is probably linked to the presence of polar nano-regions [1].

In this work we aimed to a direct visualization of polar nano-regions in paraelectric (Ba0.6Sr0.4)TiO3 phase based on oxygen atoms displacements measured from STEM images.

The chemical composition fluctuations (Ba/Sr ratios) in BST were correlated with the appearance of polar nano-regions. The approaches used we described in recently published work where the concentration of bismuth vacancies at domain walls in BiFeO3 were studied [2]. The methodology, error estimation and results obtained from image simulations and experimental annular bright-field (ABF) and high-angle annular dark-field (HAADF) images, acquired with Cs probe-corrected STEM will be explained and discussed.

[1] Hashemizadeh, et al., Journal of Applied Physics, 119, 094105 (2016) [2] Rojac, et al., Nature Materials 16, 322 (2017)

Figure 1: a.) ABF micrograph of BST in [110] zone, b.) oxygen vs Ti columns displacements indicating the polarization, c.) intensity ratio distribution of A columns (Ba/Sr ratio)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-I1

Session: Materials Science and Energy Materials Pressure Induced Effects During in situ Characterization of Supported Metal Catalysts Monia R. Nielsen*1, Jakob B. Wagner1, Christian D. Damsgaard1,2, Max Schumann3, Anker D. Jensen3, Jakob M. Christensen3 and Thomas W. Hansen1 1Center for Electron Nanoscopy,

Technical University of Denmark, 2800 Kgs. Lyngby, Denmark of Physics, Technical University of Denmark, 2800 Kgs. Lyngby, 3 Denmark Department of Chemical and Biochemical Engineering, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark *E-mail: moniarn@dtu.dk 2Department

Keywords: catalysis, pressure effects in closed cell, in situ characterization, dynamics, ETEM. In situ TEM were used to investigate dynamic processes of supported metal catalysts, such as Rh-based catalysts for the synthesis of higher alcohols, where the pressure was varied between a few millibars and atmospheric pressure.

Supported Rh-based catalysts have been widely studied for the synthesis of ethanol and other C2+ oxygenates from syngas and show promising activity and selectivity. It is therefore considered as one of the better materials for converting syngas directly into ethanol [1].

Under these in situ investigations of Rh-based catalysts, we aim to look into, among other things, strong metal-support interactions (SMSI), see Fig. 1, and to demonstrate the changes that occur at different pressures, which is an important result of the influence of pressure in terms of obtaining the structure-activity correlation for nanoparticles in reaction conditions.

First, we look at the SMSI effect at low pressure (a few millibars), see Fig. 1, using a CScorrected FEI Titan 80-300 environmental TEM and a DENSsolutions Wildfire S3 MEMS based heating holder. In order to further understand what happens at higher pressure, a DENSsolutions Climate system was used, where the catalyst can be exposed to the same gas composition at pressures up to 1 bar.

Furthermore, since the high induced electron energies and current densities typically used in HRTEM measurements often causes beam induced dynamic processes, we want to address some of the effects that occur at higher pressures.

Figure 1: Rh particles on a TiO2 support in 3 mbar H 2 illustrating the SMSI effect.

[1] V. Subramani and S.K. Gangwal, Energy & Fuels 22, 814–839 (2008).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-O1

Session: Materials Science and Energy Materials Direct Atomic-Scale Observation of Droplets Coalescence Driven Nucleation and Growth of Supported Bismuth Nanocrystal in the TEM Francis Leonard Deepak* and Junjie Li Nanostructured Materials Group, Department of Advanced Electron Microscopy, Imaging and Spectroscopy, International Iberian Nanotechnology Laboratory (INL), Avenida Mestre Jose Veiga Braga 4715-330, Portugal. *E-mail: leonard.francis@inl.int Keywords: dynamic processes, liquid/solid interface, Bi/SrBi2Ta2O9, in-situ atomic-scale observations, aberration corrected transmission electron microscopy.

Unravelling dynamical processes of liquid droplets at liquid/solid interfaces and the interfacial ordering is critical to understanding solidification, liquid-phase epitaxial growth, wetting, liquid-phase joining, crystal growth, and lubrication process, all of which are linked to different important applications in material science [1,2]. Many studies have been reported with the indirect evidence of density fluctuations at liquid/solid interfaces on the basis of X-ray scattering methods, atomic force microscopy (AFM) and with the support of atomistic simulations [3,4].

In this work, we observe in-situ atomic-scale behavior of Bi droplets segregated on SrBi2Ta2O9 by using aberration corrected transmission electron microscopy and demonstrate ordered interface and surface structures for the droplets on the oxide at the atomic-scale and unravel a nucleation mechanism involving droplet coalescence at the liquid/solid interface (Fig. 1). We identify a critical diameter of the formed nanocrystal in stabilizing the crystalline phase and reveal lattice induced fast crystallization of the droplet at the initial stage of the coalescence of nanocrystal with droplet. Further sequential observations show the stepped coalescence and growth mechanism of the nanocrystals at the atomic-scale. These results offer insights into the dynamic process at liquid/solid interfaces, which may have implications for many functionalities of materials and their applications [5].

Figure 1: Coalescence induced crystallization (Droplet 1 and 2) at the Bi/SrBi2Ta2O9 interface. The electron dose rate is 4.22 × 104 e-/Å2•s.

[1] S.H. Oh, Y. Kauffmann, C. Scheu, W.D.Kaplan, and M. Rühle, Science 310, 661-663 (2005).

[2] O.G. Shpyrko, R. Streitel, V.S. Balagurusamy, A.Y. Grigoriev, M. Deutsch, B.M. Ocko, M. Meron, B. Lin, and P.S. Pershan, Science 313, 77-80 (2006).

[3] H. Reichert, O. Klein, H. Dosch, M. Denk, V. Honkimäki, T. Lippmann and G. Reiter, Nature 408, 839-841 (2000).

[4] S. Biggs, P. Mulvaney, J. Chem. Phys. 100, 8501-8505 (1994).

[5] J. Li, Z. Wang, F.L. Deepak, ACS Nano 11, 5590-5597 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-O2

Session: Materials Science and Energy Materials Nanoscale Characterisation of the Adhesion Mechanism in Thin Metal Films for Plasmonic Applications Mario Frederik Heinig*1, Matteo Todeschini1, Johneph Sukham2, Radu Malureanu2, Alice Bastos da Silva Fanta1, Henri Jansen1, Jakob Birkedal Wagner1, Shima Kadkhodazadeh1 1

2

DTU Danchip/Cen, Technical University of Denmark, Kgs. Lyngby, 2800, Denmark Department of Photonics Engineering, Technical University of Denmark, Kgs. Lyngby 2800, Denmark *E-mail: marhein@dtu.dk Keywords: adhesion layer, metal thin film.

Deposition of thin metal films on dielectric or semiconductor substrates is central to many technological applications, including plasmonics and microelectronic devices. In this respect, good adhesion between the deposited metal and the underlying substrate is necessary, in order to ensure device integrity and performance. For plasmonic applications, noble metals such as gold and silver are the most popular choices. However, achieving the required characteristics of ultra-thin and ultra-smooth layers for plasmonic waveguides and hyperbolic metamaterials is a challenge. Gold, while more chemically stable than silver, exhibits poor adhesion to underlying substrates, requiring the deposition of a second material in between (adhesion layer), in order to obtain uniform coverage. Here, we investigate the adhesion mechanism between gold and silicon oxide substrates, using the typically chosen Cr and Ti adhesion layers, as well as organosilane adhesion layers. High-resolution transmission electron microscopy (HRTEM), transmission Kikuchi diffraction (TKD) and electron energy-loss spectroscopy (EELS) are used to understand and compare the morphology, nanostructure and chemistry of the thin film structures. The results are examined with respect to the optical properties of the corresponding structures.

Figure 1: HAADF STEM images of a 10 nm Au layer deposited on SiO2 substrate using different adhesion layers in cross-sectional geometry.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P1

Session: Materials Science and Energy Materials Regeneration of Sulfur-Poisoned Pd/Al2O3 Catalysts Mari Honkanen*1, Marja Kärkkäinen2, Mika Huuhtanen2, Hua Jiang3, Kauko Kallinen4, Riitta L. Keiski2 and Minnamari Vippola1.

1

Laboratory of Materials Science, Tampere University of Technology, Tampere, Finland.

2 Environmental and Chemical Engineering, University of Oulu, Oulu, Finland.

3 Department of Applied Physics, Aalto University, Espoo, Finland.

4 Dinex Ecocat Oy, Vihtavuori, Finland. *E-mail: mari.honkanen@tut.fi

Keywords: Pd-based catalyst, S-poisoning, regeneration, transmission electron microscopy.

The use of natural gas (NG) as an energy source will reduce the NOx and PM emissions from transportation. However, exhaust gases from NG vehicles contain unburned methane, which has to be converted to less harmful compounds. Problems related to thermal aging and poisoning occur over e.g. Pd/Al2O3 catalysts. Thermal deactivation causes the loss of active surface area. Poisoning is due to the adsorption of impurities on the catalytically active sites; a small amount of SO2 is enough to poison the catalyst. SOx compounds can be decomposed from the catalyst by regeneration treatments and thus catalyst lifetime can be extended.

In this work, regeneration of a sulfur-poisoned Pd/Al2O3 oxidation catalyst was studied.

The catalysts were characterized by (S)TEM, XRD, FT-IR, and performance tests. During the S-treatment, PdO crystallites grew slightly and bulk Al2(SO4)3 was formed. In addition, the performance, i.e. CH4 oxidation, of the S-treated catalyst decreased significantly compared to the fresh catalyst. In the regeneration treatment with CH4, a part of bulk Al2(SO4)3 was removed from the catalyst and PdO particles were partly reduced to metallic Pd. In addition, sulfur species on the Pd particles were observed due to the migration of sulfate species from the Al2O3 support onto the Pd particles. The catalyst performance was restored partly. The catalyst was further regenerated in air to reoxidize Pd particles. After this, the catalyst had still a small amount of bulk Al2(SO4)3 but sulfur species on the Pd particles disappeared. The oxidative treatment was found to have no significant effect on the catalyst performance compared to the CH4-regenerated catalyst. Thus, it can be concluded that even a small amount of bulk Al2(SO4)3 and sulfur species on the Pd have a stronger effect on the catalyst’s CH4 oxidation activity than the state of palladium (Pd/PdO) [1]. (a)

(b)

(c) PdO

PdO

Pd

Figure 1: BF STEM images of the palladium particles in the (a) fresh catalyst, (b) Spoisoned catalyst, and (c) CH4-regenerated catalyst.

[1] M. Honkanen, J. Wang, M. Kärkkäinen, M. Huuhtanen, H. Jiang, K. Kallinen, R.L. Keiski, J. Akola and M. Vippola, Journal of Catalysis 358, 253–265 (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P2

Session: Materials Science and Energy Materials Characterisation of Beam Sensitive Quartz by Scanning TEM Jochen Busam1, Sigurd Wenner2, Astrid-Marie F. Muggerud3 and Antonius T. J. van Helvoort*4.

1

Department of Materials Science and Engineering, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.

2 . SINTEF Industry, Trondheim, Norway.

3 . The Quartz Corp, Drag, Norway.

4 Department of Physics, NTNU, Trondheim, Norway. *E-mail: a.helvoort@ntnu.no Keywords: Natural quartz, dose, scanning precession electron diffraction, beam damage

Quartz is of industrial importance and extensive research has been done on it, but there are still open fundamental questions. Transmission electron microscopy (TEM) on quartz however, has hardly been published recently. The reason for this, beside challenges in preparing good specimens, is that this mineral is very beam sensitive and rapidly becomes amorphous during TEM investigations. Here, we present an assessment of modern fieldemission gun (FEG) TEM (at 200 kV) for the study of high purity quartz. To get large areas to study and avoid radiation damage during preparation, bulk specimens of up to 3000 µm2 of electron transparent material were prepared solely by mechanical wedge polishing from a petrographic thin section. Specimen charging was avoided by either using sand specimens with >1 µm grain size on a C-support or low-current scanning techniques.

One parameter affecting the rate of amorphisation is the amount of adjacent crystal to the illuminated volume. Scanning techniques with a finer probe size thus have a higher critical dose. For the scanning TEM (STEM) mode in a double corrected cold FEG JEOL ARM200F, a critical dose of 10.6 ± 2.6 [1024 e/m2] was found for a probe size <0.8 Å, a 4 pA probe current, a specimen thickness 80 nm and a [001] orientation. Both annular bright-field and annular dark-field STEM lattice imaging can be achieved. Using non-rigid registration on a series of very low dose ABF frames as well as periodical and rotational averaging [1] it was possible for the first time to resolve single Si and O atom columns in beam sensitive quartz.

Beam damage can manifest itself as strain centres with an amorphous core, which were found to grow continuously. Under repeated STEM exposure, strain centres induced by TEM illumination expanded. An alternative route to study quartz at low dose is by scanning precession electron diffraction (SPED) on an uncorrected Schottky FEG. Several SPED scans could be applied without visually changing the specimen. Post-acquisition data analysis of the SPED data allows to visualize the strain around the centres. It was also possible to acquire SPED data on grain boundaries and dislocation networks. Furthermore, the low dose SPED method allowed to study the β- to α-quartz transition at 572.5°C using a DENS heating holder. The whole grain changed phase instantaneously and no intermediate phase or twinning as previously been reported by [2] was observed. With continuous irradiation, the temperature of the β- to α-quartz was found to slightly decrease.

[1]. S. Wenner et al, Micron 96 (2017) 103–111.

[2]. G. van Tendeloo, J. van Landuyt, S. Amelinckx, Phys. Stat. Sol. 33 (1976) 723–735.

[3]. The authors thank the Norwegian Research Council and industrial partners for support to Centres for Environment-friendly Energy Research (FME) and NORTEM (grant 197405).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P3

7

Session: Materials Science and Energy Materials Microstructural Characteristics of Fretting Scars Verner Nurmi1, Jouko Hintikka2, Mari Honkanen1, Janne Juoksukangas1, Antti M채ntyl채2, Tero Frondelius2, Arto Lehtovaara1, Minnamari Vippola*1 1

Tampere University of Technology, Laboratory of Materials Science, Tampere, Finland.

2 W채rtsil채 Finland Oy, Vaasa, Finland. *E-mail: minnamari.vippola@tut.fi Keywords: fretting, SEM, EBSD, plastic deformation, fretting surface layers.

Fretting movement is dangerous for machines and machine parts because it can cause cracking and surface degradation, i.e., fretting wear and fatigue. The purpose of this work was to study fretting test pieces from the material scientific point of view: to find out what happens in the material during fretting. This work concentrates on the test pieces obtained in earlier fretting studies, using the so-called annular flat-on-flat fretting device. Fretting tests were performed with low nominal normal pressures and in gross-sliding and partial slip conditions. Specimens with annular flat-on-flat contact encountered reciprocating tangential displacement without any edge effects in the direction of the fretting movement. Various size of fretting scars were found on the surfaces of 34CrNiMo6 QT steel pairs. Main characterization tools used to study the fretting specimen cross-sections were an optical microscope, Vickers hardness test device, and SEM-EDS. EBSD maps were also created to study plastic deformation under the fretting scar, degradation layers, and interfaces between them.

Severe plastic deformation took place in fretting scar areas, deformation level being higher near the contact surface. Plastic deformation and grain orientation occurred in the direction of initiated cracks. Formation of the tribologically transformed structure was also observed in the immediate presence of fretting scar. Oxidized third body layer was found in gross-sliding samples, usually as relatively thin layers. It can be concluded, that in fretting scars various kind of material degradation takes place (Fig. 1).

Figure 1: (A) Image of fretting scar from contact surface (scale bar 1 mm). (B) Crosssectional SEM image from fretting scar showing degradation layers (scale bar 200 mm). (C) EBSD map showing plastic deformation layer under degradation layer (scale bar 20 mm).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P4

Session: (Materials Science and Energy Materials) Transmission Electron Microscopy Studies of Manganese Perovskite Electrodes for Electrochemical Water Splitting Daniel Busse*1, Janis Geppert1, Vladimir Roddatis1, Marcel Risch1 and Christian Jooss1.

1

Institute of Materials Physics, University of Goettingen, Friedrich-Hund-Platz 1, 37077 Goettingen, Germany. *E-mail: dbusse@ump.gwdg.de Keywords: perovskite oxides, TEM, electrocatalysis, oxygen evolution reaction.

Many studies have taken transition metal oxides under consideration in the search of promising oxygen evolving catalysts for electrochemical water splitting. Perovskites exhibit a wide range of chemical doping possibilities, leading to a broad spectrum of related changes in electronic structure and materials properties while keeping the perovskite crystal structure preserved. These control options allow designing catalysts for water oxidation. Several investigations have demonstrated promising catalytic activity of manganite perovskites and displayed different trends in activity and stability [1,2]. However, an in-depth understanding of active states under working conditions as well as possible pathways for corrosion is still needed.

In this contribution, epitaxially grown thin films of Pr1-xCaxMnO3 are electrochemically characterized by means of a rotating-ring disk electrode and subsequently analyzed postmortem using TEM, which provides insight into local microstructural changes and defect chemistry. Results are shown for several levels of doping (x=0.1 to 0.95), which represent a wide range of the phase diagram. EELS and HR(S)TEM have been performed to study structural, chemical and electronic changes evolved during electrochemical application. The valence state of manganese – as indicated via the O-K-edge structure as well as the Mn-L3/L2intensity ratio – has been studied . The comparison of pristine and electrochemically treated samples is discussed and linked to measurements of macroscopic electrochemistry. A decrease of the Mn-valence state in Pr1-xCaxMnO3 points to oxygen vacancy formation and suggests their participation in the oxidation mechanism. Oxygen vacancies seem to be the active sites and therefore may play an important role in many reaction steps of the OER [1].

These observations are related to proposed mechanisms of oxygen evolution and corrosion and shown in contrast to results on the catalytic activity of La1-xSrxMnO3 [3].

[1] S. Raabe, D. Mierwaldt, J. Ciston, M. Uijttewaal, H. Stein, J. Hoffmann, Y. Zhu, P. Blöchl and C. Jooss, Advanced Functional Materials 2012, 22, 3378.

[2] D. Mierwaldt, V. Roddatis, M. Risch, J. Scholz, J. Geppert, M. E. Abrishami and C. Jooss, Advanced Sustainable Systems 2017, 1, 1700109.

[3] J. Scholz, M. Risch, G. Wartner, C. Luderer, V. Roddatis and C. Jooss, Catalysts 2017, 7, 139.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P5

Session: Materials Science and Energy Materials Material properties of self-organizing aluminium nanowires in amorphous silicon Annett Thøgersen1, Torunn Kjeldstad2, Ingvild J. T. Jensen1, Marit Stange1, Ole Martin Løvvik1, and Spyros Diplas1 1 SINTEF Industry, P.O.Box 124 Blindern, 0314 Oslo, Norway 2 Department of Physics, Centre for Materials Science and Nanotechnology, University of Oslo, P.O. Box 1048 Blindern, N-0316 Oslo, Norway. *E-mail: annett.thogersen@sintef.no Keywords: Aluminium, nanowires, EELS, TEM Nanostructured materials, including nanowires, particles and tunnels, have shown to have unique optical properties compared to their bulk counterparts, like increased band gap, indirect to direct band gap transition [1], and an increased charge carrier concentration [2,3].

Such nanostructures can therefore be interesting for use in optoelectronic devices, such as solar cells and sensors. Their high specific surface area also makes them interesting for energy storage purposes.

Utilizing the immiscibility between Al and Si resulted in self-organizing Al nanowires in an amorphous silicon (aSi) matrix during magnetron co-sputtering. Removing the Al nanowires by etching created an aSi matrix with nanotunnels, similar to a honeycomb structure. However, much of the aSi has been oxidized in this process. These nanostructures may have the potential for use in Li-ion batteries [1] and photovoltaic applications.

The growth and optical properties of the Al nanowires and etched Si nanostructured films have been characterized using scanning transmission electron microscopy (STEM) and electron energy loss spectroscopy (EELS).

Figure 1: Sketch of the sample structure. A) Al nanowires in aSi, and B) The etched structure, showing aSi with nanotunnels. The figure shows the corresponding plane-view and cross-sectional images, and low loss EELS spectra of the two areas.

References [1] C. K. Chan et al. Nature Nanotechnology 3, 31-35, (2008) [2] X. Duan et al. Nature 409, 66 (2001) [3] Y. Xia et al. Advanced Materials 15, 353 (2003)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P6

21

Session: Materials Science and Energy Materials Structural and Electronic Properties of Fe Dopants in Cobalt Oxide on Au(111) : Atomistic Insight into Synergistic Effects in Mixed Metal Oxide Electrocatalysts Zhaozong Sun*1, Jonathan Rodriguez-Fernandez1, Liang Zhang2, Ting Tan2, Jakob Fester1, Aleksandra Vojvodic2, and Jeppe V. Lauritsen1.

1

Interdisciplinary Nanoscience Center (iNANO), Aarhus University, 8000 Aarhus C, Denmark 2 Department of Chemical and Biomolecular Engineering, University of Pennsylvania, Philadelphia, PA, USA * E-mail: zhaozong.sun@inano.au.dk Keywords: cobalt oxides, binary oxides, Fe dopants, scanning tunneling microscopy (STM), X-ray photoelectron spectroscopy (XPS).

Mixed metal oxides of earth-abundant 3d transition metals is an interesting class of materials that show significant synergistic effects as catalysts for electrochemical oxygen evolution compared to simple unary oxides [1, 2]. However, the exact atomic-scale nature of such mixed oxide phases and the link to their interesting chemical properties are poorly understood.

Here, a combination of scanning tunneling microscopy (STM) and x-ray photoemission spectroscopy (XPS) reveals that Fe species embed in a facile way into CoO bilayers on Au(111) resulting electronically modification of the CoO matrix. Density functional theory (DFT) modeling and fingerprint from XPS spectrum indicate the Fe dopants possess a higher oxidation state than in the structurally corresponding unary bilayer oxide. Further atomic-scale analysis suggests the substituted Fe atoms are structurally displaced further away from the substrate than the metal in either of the corresponding unary oxides. Both nearby O and Co atoms in the nearest coordination shell are geometrically and electronically perturbed. Through chemical modification and DFT calculations, high-symmetry domains are assigned to the moiréstructured nanoislands. Moiré-related properties of the Fe dopants, such as appearance and statistical distribution, are analyzed. Our results indicate that the chemical environment of neiboring O and Co atoms at the doping sites would be modified by the Fe species reciprocally and our findings may enable a better understanding of the synergistic effects of mixed metal oxides in catalytic applications. .

b)

a)

4Å

c)

4Å

Figure 1: a, c) metal/O mode STM images and b) proposed ball model of Fe doping site.

[1] Man, Isabela C., Hai-Yan Su, et al., ChemCatChem 3(7), 1159-1165 (2011).

[2] Morales-Guio, Carlos G., et al., J. Am. Chem. Soc. 138(28), 8946-8957 (2016).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P7

Session: Materials Science and Energy Materials Quantitative Microscopic Characterisation of Natural ‘Invisible’ Gold Takeshi Kasama*1, Platon N. Gamaletsos1, Stephane Escrig2, Yoshiaki Kon3, Tonci B. Zunic4, Berit Wenzell1, Louise H.S. Jensen2, Anders Meibom2, Tetsuichi Takagi3, Dimitrios Dimitriadis5, and Athanasios Godelitsas6 1

Cen/Danchip, Technical University of Denmark, Kongens Lyngby, Denmark 2 École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland 3 National Institute of Advanced Industrial Science and Technology, Tsukuba, Japan 4 University of Copenhagen, Copenhagen, Denmark 5 Hellas GOLD S.A., Athens & Chalkidiki, Greece 6 National and Kapodistrian University of Athens, Athens, Greece *E-mail: tk@cen.dtu.dk Keywords: geomaterial, precious metal, sulphide, different length scale In some gold deposits, gold exists as invisible form in pyrite and/or arsenopyrite and its concentration is extremely low. Such gold is not recovered readily from ore materials. The identification of the gold form and distribution may provide some insight into the recovery of gold. Here we use various microscopic techniques such as electron microscopy, NanoSIMS and laser ablation ICP-MS to investigate natural ‘invisible’ gold from the Olympias sulphide ore deposit, northern Greece.

Bulk analyses show that the ore samples contain gold up to ~30 ppmw and gold seems to have a positive relationship with arsenopyrite. Assuming that all the gold is incorporated into arsenopyrite, each arsenopyrite grain would contain ~200 ppmw. Laser ablation ICP-MS was used to study elemental distributions from a large region of typically 1.5 × 2 mm in steps of 50 µm. Gold is present primarily in arsenopyrite and distributes at higher concentrations towards the rim. It is clearly shown that gold has a strong correlation with arsenic. NanoSIMS measurements of arsenopyrite support the laser ablation ICP-MS results and suggest that gold is distributed with large variations in a micrometre range along the growth direction. Some gold is in pyrite; however, its concentration is even lower than that in arsenopyrite, which is also supported by SEM-WDS, where the concentration was below the detection limit (~80 ppmw) in the experimental conditions used. On the other hand, gold in arsenopyrite is detectable and its concentration is a few hundreds of ppmw, which is in good agreement with the estimate from the bulk analyses. TEM was used to examine the near-surface of arsenopyrite, where gold is usually concentrated. However, no gold precipitates were observed, which might suggest that gold is present in the form of ions in either the crystal lattice or interstitial position. This result is also supported by a previous study of a similar sample using synchrotron radiation XANES spectroscopy, in which gold is likely present as a chemically bound form in the structure of arsenopyrite [1].

As shown in the present study, the combination of these microscopic/spectroscopic techniques is very powerful to detect localised elements quantitatively and covers over a wide range of length scales. Especially it would be very useful for exploring natural mineral resources and environmentally concerned elements occurring at nanometre scale.

[1] A. Godelitsas, E. Tzamos, A. Filippidis, D. Sokaras, T.-C. Weng, G. Griego, A. Papadopoulos, S. Stoulos, P.N. Gamaletsos, T.J. Mertzimekis, E. Daftsis, and D. Dimitriadis, V.M. Goldschmidt Conference abstract, 1062 (2015).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P8

Session: Materials Science and Energy Materials

2

Microscopic investigations on the oxidation of silicon containing FeCrAl alloys V. Asokan*, A. Persdotter, J. Eklund, S. Bigdeli, T. Jonsson Environmental Inorganic Chemistry, Department of Chemistry and Chemical Engineering, Chalmers University of Technology, SE-412 96, Göteborg, Sweden *Email: asokan@chalmers.se Keywords: High temperature oxidation, FeCrAl alloys, Silicon, focused ion beam, scanning transmission electron microscopy The present study investigates the effect of silicon (Si) on the properties of the thin oxides on FeCrAl alloys using focused ion beam – scanning electron microscopy (FIB-SEM), high-angle annular dark field imaging in scanning transmission electron microscopy (HAADF-STEM) and energy dispersive X-ray spectroscopy (EDS). FeCrAl alloys are commonly used in high temperature applications because of their ability to form a protective α-alumina. However, at medium high temperatures, transient alumina may instead form and tend to contain varying amounts of Fe and Cr, depending on the temperature and composition. The corrosion behavior of FeCrAl alloys at medium high temperature is relatively unknown. Minor addition of Si to FeCrAl alloys has been shown to improve the oxidation behavior, in particular, the growth rate of the iron-rich oxide is significantly reduced.

To investigate the effect of Si on protection of FeCrAl alloys against oxidation, three samples with varying Si content have been produced and isothermally exposed in a horizontal tube furnace at 600°C in 5% O2 + N2(bal.) for 168 hours to form thin protective oxides. Imaging of sample in secondary electron mode and energy-selective backscattered mode of SEM (Ultra55 Zeiss) provided information about the oxide layer formation on the surface. In addition to the thin oxide layers, outward grown thick oxide layers are identified in FeCrAl alloy without Si, while in 1%Si and 2%Si containing FeCrAl alloy samples, almost uniformly formed thin layers are found. To characterize the oxidized regions in TEM, dual beam FIB-SEM (FEI Versa 3D) is employed to lift out thin cross-sectional region covering thin and thick oxides in the bulk specimen. Protective platinum layer is coated on the chosen region and after milling down the surrounding area, thin cross-sectioned samples are fixed to Cu TEM grid. Imaging of inward and outward growth of oxide layers are done using FEI Titan 80-300, operated at 300kV in HAADF-STEM mode. The qualitative analyses on the elemental composition of oxide regions are calculated from the EDS spectra.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P9

24

Session: Material Science and Energy Materials.

Carbon and oxygen in electrodeposited Fe coatings J. O. Nielsen*1, F. Grumsen1, J. B. Wagner2, P. Møller1, and K. Pantleon1.

1. Technical University of Denmark, Department of Mechanical Engineering, 2800 Lyngby 2. Technical University of Denmark, Center of Electron Nanoscopy, 2800 Lyngby *E-mail: jacobon@mek.dtu.dk Keywords: CBED, nanocrystalline, X-ray diffraction, Electron microscopy.

Electrochemical deposition of iron-carbon (Fe-C) coatings are a promising alternative to the banned hard chrome coatings, as they allow environmental friendly deposition and provide excellent mechanical properties. The Fe-C coating can be deposited up to several hundred microns using non-toxic chemicals at 50 ℃. The nanocrystalline coatings with grain sizes in the order of 10-20 nm provide high hardness of around 780 to 800 HV. The coating has a strong fibre texture with a preferred <311> orientation and columnar grains in the growth direction. A thorough material characterization of the Fe-C coating is essential for understanding the film growth and tailoring the resulting mechanical properties.

A chemical analysis of the Fe-C coating revealed a carbon concentration of 0.85 wt%, together with some oxygen and hydrogen as byproducts from the deposition process. Despite the very low solubility of both carbon and oxygen in ferrite, corresponding oxide and/or carbide phases have not been identified unambiguously. Phase identification by means of conventional X-ray diffraction (XRD), synchrotron diffraction, selected area electron diffraction (SAED) and convergent beam electron diffraction (CBED) mainly revealed the presence of ferrite. Because of several experimental challenges and limitations for each of the applied characterization methods, a complementary analysis of the chemical composition, and the size and orientation of grains is applied to identify possible carbides (e.g. η-Fe2C) and oxides (e.g. Fe3O4) in the Fe-C coatings. Alternatively to the formation of carbide and oxide phases, significant supersaturation of the ferrite phase with carbon and oxygen may result from the electrodeposition process, which is known to cause deviations from thermodynamic equilibrium. However, neither bct martensite, corresponding to supersaturation of ferrite with carbon, nor any changes of the lattice spacing of bcc ferrite due to solid solution of oxygen or carbon, have been detected. An increase in oxygen concentration was found in the grain boundaries using scanning transmission electron microscopy (STEM) and electron energy loss spectroscopy (EELS) by measuring in plane-view of the columnar microstructure. Atom probe tomography (APT) further supplemented information on the chemical distribution of carbon and oxygen in the coating.

Based on the complementary results from spectroscopic, microscopic and diffraction techniques, the internal structure of the Fe-C coatings is discussed.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P10

Session: Materials Science and Energy Materials Electron microscopy as a means to validate Raman spectroscopy for quantifying single-walled carbon nanotubes Hua Jiang*1, Ying Tian1,2, and Esko I. Kauppinen1.

1

2

Department of Physics, Aalto University School of Science, Espoo, Finland.

Department of Physics, Dalian Maritime University, Dalian, Liaoning 116026, China. *E-mail: hua.jiang@aalto.fi

Keywords: electron diffraction, Raman spectroscopy, SWCNTs, quantitative analysis.

Recently there is a growing trend of using resonance Raman technique to quantify the population of a specific type of conductivity in a bulk material. In this work, we use electron diffraction as a means to evaluate its validity. Three SWNTs samples with different diameter distributions, including a ferrocene decomposition floating catalyst chemical vapor deposition (ferrocene-FC-CVD) SWNT sample [1], and a spark-based FC-CVD (spark-FC-CVD) sample [2], in addition to a NIST SWNT reference sample (RM8281), have been investigated. Raman spectroscopy study with three excitation wavelengths of 514, 633, and 785nm was performed to quantify the metallic SWNT concentrations (M%) in the samples. To evaluate the Raman results, electron diffraction (ED) technique was used to directly map the chirality distribution.

In the ferrocene-FC-CVD sample, M% was estimated over 90% from Raman analysis at 633nm, but 0% at 514nm, while ED analysis gave about 24%. For the spark-FC-CVD sample, the 633nm Raman analysis led to about 55% metallic tubes, but the 514 laser resulted in less than 2% though ED analysis turned out to be 33%. In particular, the Raman assessment of the well-known (6,5)-dominated NIST reference sample at all three wavelengths of 633nm, 514nm and 785nm, however, showed a small minority of (6,5) tubes, due to the weak resonance of the (6,5) tube with any of those lasers.

To conclude, our results prove that the Raman RBM intensities depend largely on the resonant conditions at certain wavelengths, rather than simply on concentrations. Up to the resonance conditions, some majority nanotube species revealed by electron diffraction measurements induce relatively weak, or even missing RBMs, and vice versa. This certainly leads to an uncertainty over Raman spectroscopy for quantitative assessment of metallic tube concentrations calculating from the relative peak intensities [3].

[1] A. Kaskela, et. al, Carbon, 103, 228 (2016).

[2] K. Mustonen, et. al, Appl. Phys. Lett., 107, 013106 107(2015).

[3] Y. Tian, et. al, Anal. Chem. 90, 2517 (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P11

25

Session: Materials Science and Energy 3D-Orientation Mapping of Nano-grains in Solid Oxide Electrochemical Cells using TEM 1

S. Colding-Jørgensen , S. B. Simonsen1, S. Schmidt2, W. Zhang1 and L. Theil Kuhn1.

1

2

DTU Energy, Technical University of Denmark, Risø. Denmark DTU Physics, Technical University of Denmark, Kgs. Lyngby. Denmark *E-mail: sofcol@dtu.com

Keywords: Nano-grains, Orientation mapping, SOEC, 3D-OMiTEM, TEM. The performance of functional materials such as solid oxide fuel cells (SOFC) is closely related to the nano- and micro granular structure due to the large fraction of grain boundaries [1]. Orientation effects on conductivity in SOFCs have previously been studied using impedance spectroscopy [1][2][3]. This showed that by aligning the particles, the ionic conductivity can be enhanced by up to two orders of magnitude in 2D thin films. However, 3D bulk samples appear to have different electrical properties and the experimental results point in different directions. Researchers discuss the validity of the experimental impedance results since cracks and other 3D structural defects can affect the measurement. Impedance measurements should therefore be accompanied with a visualization of the 3D structure. In order to successfully understand the relationship between the performance and structure of SOFCs, it is thus necessary to visualize them nondestructively in 3D. We propose 3D Orientation Mapping in TEM (3D-OMiTEM) [4], a promising technique for non-destructive visualization in 3D. A feasibility study on aluminum showed that 3DOMiTEM can be done on approximately 100 nm thick samples with grains smaller than 20-30 nm with a resolution of a few nm. 3D-OMiTEM utilizes dark-field conical scanning in combination with sample tilts to obtain combined reciprocal and direct space information.

The conical scanning uses beam tilt to choose diffraction rings from which dark field images are formed. Therefore, the reciprocal information is given by the beam tilt and sample rotation. Each measurement thus constitute a 2D-projection of the 3D grain shapes observed for a point in the reciprocal space. The subsequent reconstruction procedure provides orientation determination locally in the interior of the sample. Typically, each of the 6-10 inner rings in the diffraction pattern are used for every sample rotation angle. This results in up to 10.000 images that are reconstructed into a 3D grain orientation map.

Here we apply the 3D-OMiTEM method on 8% Yttria-stabilized Zirconia (8YSZ) which is a state of the art material for SOFCs. Electro spun and calcined 8YSZ nanofibers with the high ion conducting cubic phase were chosen since they have constant thickness when rotated in 3D-OMiTEM. The talk will give an overview of 3D-OMiTEM studies of YSZ. [1] S. H. Jo, P. Muralidharan and D. K. Kim. Solid State Ionics, 178, 1990–1997 (2008) [2] G. Baure, H. Zhou, C.-C Chung, M. A. Stozhkova, J. L. Jones and J. C. Nino, Acta Materialia, 133, 81–89 (2017) [3] H. Fricke, Journal of Physical Chemistry, 57, 934–937. (1954) [4] H. Liu, S. Schmidt, H.F. Poulsen, A. Godfrey, Z. Q. Liu, J. A. Sharon, and X. Huang, Science, 332, 833–834.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P12

2

Session: Materials Science and Energy Materials Scanning Transmission Electron Microscopy Of Single Atoms And Dimers From Cluster Deposition Niklas Mørch Secher*1, Jakob Kibsgaard1, and I. Chorkendorff1 1

Technical University of Denmark, Anker Engelundsvej 1, 2800 Kongens Lyngby, Denmark *E-mail: niklas@fysik.dtu.dk Keywords: single atoms, STEM, cluster deposition, catalysis, model catalysts.

Improving existing heterogeneous catalysts requires novel understanding of the dynamics of the chemical reaction at an atomic scale. Existing catalysts have several properties that inhibit optimal characterization and understanding such as bulky geometries and a variety of active sites. Therefore, there is a need for simple model catalysts, which are more suitable for characterizing and understanding.

Such model catalysts can be created using the Nanobeam cluster source seen in Fig. 1.

Here a magnetron sputter head sputters single atoms off a metal target that can then agglomerate to a certain size. A time of flight mass filter then filters out entities with a certain mass, which are deposited on a chosen substrate. This can be used to deposit single atoms, dimers or small clusters all the way up to nanoparticles on a variety of suitable support structures.

Recently there has been a large interest in single atom catalysts since these are predicted to exhibit novel catalytic properties for some reactions of interest [1]. However, checking that these catalysts are indeed single atomic is only possible with very few techniques. One of these is Scanning Transmission Electron Microscopy (STEM) [2, 3]. Here I present STEM images of platinum single atoms (Fig. 2) and dimers deposited on a graphene sheet using the Nanobeam cluster source. Furthermore, I elaborate on some of the methods that can be employed to characterize such samples and give valuable information on their atomic structure.

Figure 1: Schematic drawing of the cluster source at DTU Physics, used for depositing single atoms and small cluster of different metals.

Figure 2: STEM image of single platinum atoms on a monolayer graphene sheet.

[1] K. Jiang et al., Energy & Environmental Science (2018).

[2] H. Chung et al., Science 357, 479-484 (2017).

[3] O. L. Krivanek et al., Low Voltage Electron Microscopy: Principles and Applications, 1, 119-161 (2013).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P13

Session: Materials Science and Energy Materials Transmission Kikuchi Diffraction characterization of low dimensional materials Alice Bastos S. Fanta1, Svetlana Alekseeva4, Christoph Langhammer4, Beniamino Iandolo3, Dmitrii Viazmitinov2, Takeshi Kasama1, Elizaveta Semenova2, Andrew Burrows1 1. DTU Danchip/Cen, Technical University of Denmark, Kgs. Lyngby, Denmark 2. DTU Fotonik, Department of Photonics Engineering, Technical University of Denmark, Kgs. Lyngby, Denmark 3. DTU Nanotech, Institute for micro and nanotechnology, Technical University of Denmark, Kgs. Lyngby, Denmark 4. Department of Physics, Chalmers University of Technology, GĂśteborg , Sweden Keywords: Transmission Kikuchi Diffraction, nanoplasmonic particles, II-V nanowires Since the first publication from Keller and Geiss [1] on transmission Kikuchi diffraction in the SEM, the technique has attracted considerable attention and has been applied to a variety of materials and samples [2]. A significant amount of this attention has been dedicated to complementing EBSD characterization with the improved spatial resolution provided by TKD, and to this end electron transparent samples from bulk materials were prepared. The characterization of low dimension materials such as, nanoparticles, nanowires and thin films, in the SEM using TKD has, however, not yet been well explored, but it has a large potential.

Many samples, which have traditionally been investigated only with the TEM, due to their resolution requirements or their small volume or thickness, can now also be investigated by TKD in the SEM. Consequently, quantitative microstructural characterization over relatively large areas and with a spatial resolution in order of 2-10 nm [3] can be achieved. This potential is demonstrated in this presentation by means of two examples. In the first example the characterization of III-V nanowires by TKD is compared to TEM results and reveals a fast approach of statistical characterization of epitaxially grown nanowires, where, for instance, phase control of the growth process can easily be investigated with statistical relevance. In the second example, nanoplasmonic particles are characterized by TKD, where a correlation between individual particles microstructure and their hydride-formation pressure is presented, and the role of grain boundaries is revealed[4].

Considering that SEMs are more commonly accessible and relatively simpler to operate compared to a TEM, these examples clearly show that characterization of low dimensional material by TKD has a large prospective and can play an important role in many fields of functionalised low dimensional materials.

Figure 1: a) TKD orientation map of InP nanowires b) single particle isotherms with TEM and TKD micrographs[4].

[1]

[2]

R. r Keller, R. h Geiss, Transmission EBSD from 10 nm domains in a scanning electron microscope, J. Microsc. 245 (2012) 245â&#x20AC;&#x201C;251. doi:10.1111/j.13652818.2011.03566.x.

G.C. Sneddon, P.W. Trimby, J.M. Cairney, Transmission Kikuchi diffraction in a

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P14

Session: Materials Science and Energy Materials

[3]

[4]

scanning electron microscope: A review, Mater. Sci. Eng. R Reports. 110 (2016) 1â&#x20AC;&#x201C;12. doi:10.1016/j.mser.2016.10.001.

P.W. Trimby, Y. Cao, Z. Chen, S. Han, K.J. Hemker, J. Lian, X. Liao, P. Rottmann, S. Samudrala, J. Sun, J.T. Wang, J. Wheeler, J.M. Cairney, Characterizing deformed ultrafine-grained and nanocrystalline materials using transmission Kikuchi diffraction in a scanning electron microscope, Acta Mater. 62 (2014) 69â&#x20AC;&#x201C;80. doi:10.1016/j.actamat.2013.09.026.

S. Alekseeva, A.B. da S. Fanta, B. Iandolo, T.J. Antosiewicz, F.A.A. Nugroho, J.B. Wagner, A. Burrows, V.P. Zhdanov, C. Langhammer, Grain boundary mediated hydriding phase transformations in individual polycrystalline metal nanoparticles, Nat. Commun. 8 (2017) 1084. doi:10.1038/s41467-017-00879-9.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P14

29

Session: Materials Science and Energy Materials Quantification of the Sigma Phase in Super Duplex Stainless Steel

by Scanning Electron Microscopy.

Jannicke Kleppen1, Ida Westermann1, Morten Karlsen1, Yingda Yu1, Torkjell Breivik2, Bjørn Eske Sørensen2, Jarle Hjelen*1,2.

1

Department of Materials Science and Engineering, NTNU, Alfred Getz vei 2, 7491 Trondheim, Norway.

2 Department of Geoscience and Petroleum, NTNU, Sem Sælands vei, 7491 Trondheim, Norway *E-mail: Jarle.Hjelen@ntnu.no

Keywords: sigma phase, stainless steel, quantification, SEM, EBSD. Super Duplex Stainless Steels (SDSS) exhibit high mechanical and corrosion resistance properties. The high alloying content in these steels increases the risk of precipitation of intermetallic phases, with a negative effect on the corrosion resistance and ductility. During heat treatment like welding the undesired intermetallic sigma phase may form. To construct the time-temperature-transformation (TTT) diagram of a particular SDSS, it is necessary to detect the first sigma phase to form and to quantify the fraction of the sigma phase as a function of heat treatment time.

Atomic number contrast imaging could be used to identify and quantify the sigma phase [1].

Another approach is to apply Electron BackScatter Diffraction (EBSD) for detection of the sigma phase. In case of Image Quality (IQ) maps [2] the sigma phase will be dark compared to the austenite and ferrite phases. Areas with topography will also be dark in conventional IQ maps, see Figure 1. However, by applying auto brightness and pattern averaging in the NORDIF EBSD 3.0 acquisition software combined with dynamic background subtraction during indexing with TSL/OIM the IQ maps will be improved and less sensitive to topography, see Figure 2 which shows the same area as in Figure 1.

An alternative to use IQ maps for detection and quantification of the sigma phase, is to apply phase maps. Even the phases have different crystal structures it is hard to get reliable patterns from the sigma phase. In areas with topography, like grain boundaries, patterns from ausenite and ferrite are often misindexed as the sigma phase. It is a challenge to optimize the SEM-EBSD parameters to get high pattern quality of the sigma phase. If the specimen preparation is unsufficient a very high fraction of sigma is detected due to misindexing.

Figure 1: Conventional IQ map.

Figure 2: Improved IQ map

[1] A. Hosseini et al. Weld World (2018).

[2] Wright, S., and Nowell, M. Microscopy and Microanalysis, 12(1), 72-84. (2006).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P15

Session: Materials Science and Energy Materials Electron Tomography of Self-Assembled Metal Nanoparticle Superstructures Nonappa*1,2, Peter Engelhardt1, and Olli Ikkala1,2.

1

Department of Applied Physics, Aalto University School of Science, Puumiehenkuja 2, FI02150, Espoo, Finland.

2 Department of Bioproducts and Biosystems, Aalto University School of Chemical Engineering, Kemistintie 1, FI-02150, Espoo, Finland. *E-mail: nonappa@aalto.fi Keywords: electron tomography, metal nanoparticles, self-assembly, colloids.

Abstract: Electron tomography (ET) and single particle reconstruction (SPR) methods have emerged as powerful tools to study three dimensional (3D) structural details of non-crystalline biological assemblies at atomic resolution [1,2]. Therefore, ET reconstructions are complementary as well as alternative tools to X-ray crystallographic studies. However, the application of ET reconstruction and SPR methods in material science, especially for metal nanoparticles is limited, partly due to polydispersity of synthetic materials. We have designed a conceptually simple, self-assembled colloidal capsid using atomically precise gold nanoclusters containing exact number metal atoms and surface ligands [3]. ET reconstructions revealed that the spherical capsids are made up of monolayer thick shell. In this presentation, we will show how ET has been utilized to investigate the mechanistic details on self-assembly of colloidal capsids (Fig.1) and hybrid platonic cages encapsulating plasmonic nanoparticles [4-6].

Figure 1: a) TEM micrograph of a spherical capsid and b), 3D reconstruction of spherical capsid showing hollow interior with a multilayered shell.

[1] J. Frank, Three-dimensional Electron Microscopy of Macromolecular Assemblies, Oxford University Press, Inc. New York, 2, 1–432 (2006).

[2] P. Engelhardt, Encyclopedia of Analytical Chemistry, 6, 4948–4984 (2006).

[3] Nonappa, T. Lahtinen, J. S. Haataja, T. –R. Tero, H. Häkkinen, O. Ikkala, Angew. Chem.

Int. Ed. 55, 16035–16038 (2016).

[4] Nonappa, J. S. Haataja, J. V. I Timonen, S. Malola, P. Engelhardt, N. Houbenov, M. Lahtinen, H. Häkkinen, O. Ikkala, Angew. Chem. Int. Ed. 56, 6473–6477 (2017).

[5] Nonappa, O. Ikkala, Adv. Funct. Mater. 27, 1704328 (2017).

[6] A. Chakraborty, A. C. Fernandez, A. Som, B. Mondal, G. Natarajan, G. Paramasivam, T. Lahtinen, H. Häkkinen, Nonappa, and T. Pradeep, Angew. Chem. Int. Ed. 58, 10.1002/anie.201802420 (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P16

Session: Materials Science and Energy Materials Jacket of Prof. Elias Lönnrot, collector of Kalevala epos Krista Vajanto*1, Jenni Suomela, and Alex Snellman3.

1

Nanomicroscopy Center Aalto University, Puumiehenkuja 2, 02150 Espoo, Finland.

2 Crat Studies, University of Helsinki, Finland.

3 History, University of Helsinki, Finland. *E-mail: krista.vajanto@aalto.fi Keywords: civil uniform, frock coat, broadcloth, metal thread embroidery

Civil uniforms [1] were a phenomenon of late 19th century in Finland, part of Russia then. For example, professors wore dark blue frock coats with metal thread embroidery and goldish buttons in front.

In the focus of our case study is professor-jacket of Elias Lönnrot, the collector of Finland’s national epos Kalevala. After the death of Lönnrot (1884), the jacket was donated to the collections of National museum of Finland. A few millimeter-long fiber samples were analysed with means of microscopy (SEM, SEM-EDX, TLM and POL) to deepen the knowledge of the materials of the jacket.

Main visible fabric of the jacket is blue broadcloth, close to merino quality. In microscopy observation, the wool fibers seemed to be coated with some organic particles, that might consist of the blue color pigment and dirt. In addition, there were clear growth of mold of fibers. This might indicate manufacturing conditions of the fabric or tell about the wear history of the jacket.

Precious materials had been used to produce the palm leaf embroidery of collar and linear embroidery of sleeve ends (Fig. 1). According to our analysis, the core thread of the embroidery is silk. Around the silk core, there is a metal lamella. That is less than 0.5 mm wide, but very evenly cut that could tell about professional manufacturing. Lamella is of silver, coated with a layer of gold.

Several different fabrics were used to create the lining of the jacket. There were twill, satin and plain weave, used symmetrically but with different widths. Visually, all fabrics seemed to be very shiny and soft, in neutral or beige-grey colors. Predominant lining material was found to be cotton, while silk was used as sewing yarn. Flax was found in support fabric.

Many different materials could tell about general reusing of fabrics, or professionally made repairs or lack/avoiding of the most expensive fabric materials.

Figure 1: Metal thread embroidery of collar, gold gilded silver lamella.

[1] O. Gripenberg, Civiluniformer i Finland (1969).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P17

Session: Materials Science and Energy Materials Post-mortem Analyses of Long-term Tested SOEC Cells in Coelectrolysis Mode Yuliang Liu*1, 2, Ming Chen1, and Xiufu Sun1.

1

Department of Energy Conversion and Storage, Technical University of Denmark, DK-4000 Roskilde, Denmark 2 School of Materials Science and Engineering, Henan University of Science and Technology, 471023 Luoyang, China. *E-mail: yuliua@dtu.dk Keywords: Solid oxide electrolysis cells (SOECs), degradation, Ni/YSZ electrode, microstructure evolution.

Solid oxide electrolysis cells (SOECs) have drawn great interest recently. By high temperature electrolysis of steam or co-electrolysis of steam and CO2, electrical energy can be converted to chemical energy and stored as H2 or synthesis gas. High performance and durable operation are the keys to bring down the cost of fuel production from SOECs. Improving the understanding of degradation mechanism and optimizing the cell operation conditions are also needed to advance the SOEC technology into the market. In recent years, extensive efforts have been devoted to studies on the cause of SOEC degradation and different degradation mechanisms have been proposed [1-4].

In this work, we present results on post-mortem analysis of SOEC cells tested under different conditions. Four nominally identical cells (hereafter referred as Cell A/B/C/D) were investigated in this work. Cell A was exposed to reduction only, while Cells B/C/D were longterm tested at 800 °C under an electrolysis current density of 1 A/cm2 with different inlet gas compositions and different conversion of steam & CO2. The cell microstructure was examined using scanning electron microscopy (SEM, Zeiss Supra 35) with energy dispersive X-ray (EDS).

The most significant microstructural changes caused by long-term co-electrolysis operation are identified to be in the active Ni/YSZ electrode. An increase in porosity is detected in all the three long-term tested cells. The porosity change is most likely caused by Ni re-distribution, moving from the active electrode layer to the neighboring support layer. This is evidenced by EDS line scan results. Ni re-distribution results in loss of active triple phase boundaries (TPBs) in the active electrode layer and eventually results in the observed cell degradation. Another microstructural change is identified to be inclusions inside Ni grains. EDS results indicate that these inclusions are most likely SiO2 with a small amount of Al2O3. These microstructural changes in the Ni/YSZ electrode are quantified and are further correlated with the test conditions. A strong correlation between the detected microstructural changes and local electrode over-potential is established.

[1] A. Hauch, S. D. Ebbesen, S. H. Jensen, and M. Mogensen, J. Electrochem. Soc., 155, B1184 (2008).

[2] S. D. Ebbesen, C. Graves, A. Hauch, S. H. Jensen, and M. Mogensen, J. Electrochem.

Soc., 157, B1419 (2010).

[3] M. Chen, Y. Liu, J. J. Bentzen, W. Zhang, X. Sun, A. Hauch, Y. Tao, J. R. Bowen, and P. V. Hendriksen, J. Electrochem. Soc., 160 (8) F883-F891 (2013).

[4] P. Moçoteguy, A. Brisse, Int. J. Hydrog. Energy., 38 15887–15902 (2013).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P18

34

Session: Materials Science and Energy Materials In-situ SEM nanomechanical testing Gaurav Mohanty1,2 and Johann Michler2 1

Materials Science, Tampere University of Technology, Korkeakoulunkatu 10, FI-33720 Tampere, Finland 2 Laboratory for Mechanics of Materials and Nanostructures, Empa â&#x20AC;&#x201C; Swiss Federal Laboratories for Materials Science and Technology, Thun, Switzerland

Recent advances in materials for structural applications have pushed the need for mechanical testing at the micro and nanoscale. This has spawned a host of micron scale techniques - some directly adapted from conventional tests for small length scales like miniaturized tension and compression tests, and others like instrumented indentation and microcompression tests that are of more recent origin. Among these, nanoindentation has emerged as the most widely used and versatile technique for mechanical testing of thin films, miniaturized devices and materials at small length scales over the past two decades, partly due to ease of testing and minimal sample preparation. The addition of in-situ observation of the material deformation characteristics in the SEM and TEM enables researchers to draw conclusions on the operative deformation mechanisms, an important criterion for structural materials design.

This talk aims to introduce the emerging techniques in the field of in-situ micron scale testing with special emphasis on nanoindentation and micropillar compression in the SEM. The use of focused ion beam (FIB) is advantageous for preparing micropillars, cantilevers, and other test geometries at specific sites like grains of known orientation, grain/phase boundaries, etc. Case studies of in-situ mechanical tests will be presented to illustrate extraction of mechanical property parameters like yield strength, fracture toughness, time dependent plasticity (creep, relaxation and strain rate sensitivity) and fatigue properties at the micron scale. Recent advances like variable temperature measurements (both cryo and high temperature testing), high strain rate tests and combining micron scale mechanical tests with x-rays, Raman and EBSD will also be presented. Finally, a brief overview of the state of the art and future research directions will be discussed.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark A-P19

35

Session: Materials Science and Energy Materials Grain Boundary Mediated Hydriding Phase Transformations In Individual Polycrystalline Metal Nanoparticles Svetlana Alekseeva*1, Alice Bastos da Silva Fanta2, Beniamino Iandolo2,5, Tomasz J.

Antosiewicz1,3, Ferry Anggoro Ardy Nugroho1, Jakob B. Wagner2, Andrew Burrows2, Vladimir P. Zhdanov1,4, and Christoph Langhammer1.

1

Department of Physics, Chalmers University of Technology, Göteborg 412 96, Sweden.

2 Center for Electron Nanoscopy, Technical University of Denmark, Fysikvej, 2800 Kgs. Lyngby, Denmark.

3 Centre of New Technologies, University of Warsaw, Banacha 2c, Warsaw 02-097, Poland.

4 Boreskov Institute of Catalysis, Russian Academy of Sciences, Novosibirsk 630090, Russia.

5 Present address: Department of Microtechnology and Nanotechnology, Technical University of Denmark, Ørsteds Pl., 2800 Kgs. Lyngby, Denmark. *E-mail: svetlana.alekseeva@chalmers.se Keywords: grain boundary, palladium, single nanoparticle, plasmonic nanospectroscopy.

Grain boundaries separate crystallites in solids and influence material properties, as widely documented for bulk materials. In nanomaterials, however, investigations of grain boundaries are very challenging and just beginning. Here, we report the systematic mapping of the role of grain boundaries in the hydrogenation phase transformation in individual Pd nanoparticles.

Employing multichannel single-particle plasmonic nanospectroscopy, we observe large variation in particle-specific hydride-formation pressure, which is absent in hydride decomposition. Transmission Kikuchi diffraction suggests direct correlation between length and type of grain boundaries and hydride-formation pressure. This correlation is consistent with tensile lattice strain induced by hydrogen localized near grain boundaries as the dominant factor controlling the phase transition during hydrogen absorption. In contrast, such correlation is absent for hydride decomposition, suggesting a different phase-transition pathway. In a wider context, our experimental setup represents a powerful platform to unravel microstructure–function correlations at the individual-nanoparticle level. [1]

H2 partial pressure (mbar)

102

a

b

c

d

e

f

s1p2

101 0 102

−0.1 −0.2 ΔPI (a.u.) s2p5

101 0

−0.1 −0.2 ΔPI (a.u.)

111

Inverse pole figure plot of the growth direction of Pd 001

101

Figure 1: Single Pd particle isotherms (a, d) with corresponding TEM (b, e) and TKD (c, f) micrographs. The scale bar in TEM images is 50 nm.

[1] Alekseeva, S. et al. Grain boundary mediated hydriding phase transformations in individual polycrystalline metal nanoparticles. Nature Communications 8, 1084, (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark C-O1

36

Session: Scanning Probe Microscopy Adsorption and Diffusion of NH3 on Anatase-TiO2 (101) Kræn Christoffer Adamsen*1, Stig Koust1, Esben Leonhard Kolsbjerg2, Bjørk Hammer2, Stefan Wendt1 and Jeppe V. Lauritsen1.

1

Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, 8000 Aarhus C. 2 Department of Physics and Astronomi, Aarhus university, Ny munkegade 120, 8000 Aarhus C. *E-mail: Kraenca@inano.au.dk

Keywords: Scanning Tunneling Microscopy, Oxide surfaces, Diffusion, Ammonia.

Fundamental understanding of catalytic processes for NOx removal (Selective Catalytic reaction, SCR) is vital for improving existing catalysts and developing new. In the SCR cycle, NOx is known to react from gas-phase on adsorbed ammonia on VOx/TiO2 based catalysts [1].

Adsorption of ammonia on such oxides is therefore of great importance for fundamental understanding of NOx-removal and SCR catalysis. Here we present a fundamental study of the static and dynamic behaviour of ammonia on anatase-TiO2 (101), the predominant facet on anatase-TiO2 nanoparticles. High resolution Scanning Tunnelling Microscopy (STM) of static adsorbed ammonia molecules at room temperature, indicates a strong binding to the surface[1]. The strong binding of ammonia was further quantified by Temperature Programmed Desorption (TPD), which also shows a highly coverage dependent binding energy, indicating molecular repulsion. Molecular repulsion also shows a clear effect in largescale atomically resolved STM images and statistical analysis of intermolecular coordination supplied repulsion energies for these. Single ammonia molecule diffusion measured utilizing the high-speed Aarhus STM, show diffusion to all neighbouring sites. High- resolution temperature controlled STM movies enables us to determine the barrier of diffusion and looking into the complexity of ammonia diffusion. We conclude that molecular repulsion increases the diffusability of ammonia molecules, however if the repulsion is overcome a surprising lower energy diffusion path was found for ammonia dimers. Here they have the possibility of diffusing through a rolling effect, where ammonia can move more easily in one direction, which resembles water dimer diffusion of water dimers on Rutile-TiO2 (110)[3].

Figure 1: Overlap of two High-resolution STM images of diffusing ammonia molecules [1]Arnarson, Logi, et al. "The reaction mechanism for the SCR process on monomer V 5+ sites and the effect of modified Brønsted acidity." Physical Chemistry Chemical Physics 18.25 (2016): 17071-17080.

[2] Koust, Stig et al. NH 3 adsorption on anatase-TiO 2 (101). The Journal of Chemical Physics. 148. 124704. (2018)

[3] Matthiesen, Jesper, et al. "Formation and diffusion of water dimers on rutile TiO 2 (110)." Physical review letters 102.22 (2009): 226101.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark C-O2

Session: Materials Science and Energy Materials

37

Preparation of Aluminum Specimen with Gallium and Xenon Plasma Focused Ion Beam for Further Nano-characterization Martin Sláma*1, 2, Petr Klímek1, Manuel Bornhoefft1, Jiří Dluhoš1, Hana Tesařová1 1

2

Tescan Orsay Holding a.s., Libusina tr. 863/21, 623 00 Brno, Czech Republic.

Institute of Materials Science and Engineering, Brno University of Technology, Technicka 2, 616 00 Brno, Czech Republic *E-mail: martin.slama@tescan.com Keywords: Focused ion beam, nano-pillars, nano-characterization, mechanical testing, aluminum

Focused ion beam scanning electron microscopes (FIB-SEM) enable highly-localized and sitespecific material removal with practically no restriction on sample composition. Depending on the ion source (e.g. Ga+, Xe+), the rate of material removal differs significantly. In general, the design of Xe+ source allows using high ion beam currents that can be up to a few µA while maintaining beam quality and performance, something which is not possible with the Ga+ sources. Such high ion currents are achievable due to the fact that the Xe+ source is broader, more collimated and has higher angular intensity. In addition, Xe+ ions are more massive than Ga+ thus more atoms are ejected from the target per incident ion, which also contributes to obtain significantly higher sputtering rates. However, the most relevant feature of Xe ions for this study is their non-metallic and inert nature which prevents any chemical interaction with the target material and formation of unwanted metallic compounds that alter the original properties of the sample that is being analyzed. Furthermore, the penetration depth of Xe ions is significantly less than Ga ion for a given energy resulting in less ion implantation that could induce local changes in the crystalline structure of the sample and, in turn, lead to changes in the mechanical properties of the sample. As some materials such as Al, Cu and Ga are sensitive to Ga ions, Xe+ ion milling can be a good asset for many FIB applications that involve working with these materials. This can be considered as a significant advantage over Ga ion source FIBs especially in microanalysis and nano-mechanical characterization. A way to perform nanomechanical characterization of materials is to prepare nano-pillars to test their mechanical properties. As the Ga+ ions penetrating into the material (aluminum in our case) during the FIB milling process, they get implanted and can even weaken the material intended for testing. In contrast, it is advantageous to prepare aluminum nano-pillars with Xe+ plasma FIB since – for the reasons mentioned above – does not introduce significant changes in the material and, as a consequence, the results will be more reliable compared to those in which the pillars had been prepared with a Ga source FIB. Taking these facts into consideration, we pursue the following objectives: (1) Demonstrate the process of pillar preparation with Ga+ and Xe+ FIB and show the advantages of the wider ion beam range offered by the Xe+ plasma FIB. (2) Indicate the influence of gallium implantation on Al materials, (3) Describe the effect of gallium on properties of micron-range grains of aluminum sample treated by the ECAP method via different methods.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark C-O3

Session: Materials Science and Energy Materials

Figure 1: TEM image of gallium contamination in Aluminum. Insets showing gallium and aluminum EDX maps of the marked region.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark C-O3

Session: Materials Science and Energy Materials Characterisation of ´/´´ precipitates of Electron Beam Melted Alloy 718 using TEM methods Lisa Lautrup*1, and Peter Harlin.

1

Sandvik Materials Technology, Strategic Research & Innovation, SE-811 81 Sandviken, Sweden 2 Sandvik Materials Technology, Product Area Powder R&D, SE-811 81 Sandviken, Sweden *E-mail: lisa.lautrup@sandvik.com Keywords: additive manufacturing, Alloy 718, SAD, particle size distribution Alloy 718 is well-known for its high strength at elevated temperatures and ability to withstand corrosive environments and is readily used by off-shore and aerospace industry. The strengthening effect is caused by formation of secondary precipitates of primarily ´´ phase and to some extent ´phase. In this study, one of the available additive manufacturing processes, electron beam melting (EBM), is utilized on Alloy 718 powder [1] to see if the mechanical properties can be achieved with this manufacturing route (Figure 1A). The advantage being less material waste and the ability to achieve more complex designs. Post treatments such as annealing and hot isostatic pressing (HIP) are tested and correlated to mechanical properties.

The size and morphology of the secondary precipitates are affected by the post-treatments and show a clear dependence on mechanical properties and can therefore be used to achieve the desired material properties. Both the number density of secondary precipitates and the form factor increases the strength of the material and should ideally be combined to achieve the highest possible strengthening effect. This effect is mainly attributed to the ´´ phase precipitates as they have the elongated shape while ´ are more spherical in shape nucleating on the ´´ rods (Figure 1C). Overlap between neighboring precipitates makes data processing difficult from HAADF-STEM (Figure 1A) data and takes both ´and ´´ into account. Careful dark field (DF) imaging using the smallest selected area diffraction (SAD) aperture (Figure 1B) can distinguish between the two precipitate types and shows the difference in particle shape (Figure 1C).

A

B

C

Figure 1: Elongated inclusions of ´and ´´ phase In Alloy 718 after EBM (A) with a given orientation relationship to the austenite () matrix (B) giving the precipitation hardening effect of Alloy 718. The ´´ (orange) are elongated and ´(blue) more spherical in shape (C).

[1] M.M. Kirka et al., Mat. Sci. Eng. A 680, 338-346 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark C-O4

Materials Science and Energy Materials Elevated Temperature In-situ Transmission Kikuchi Diffraction: a Powerful Tool for the Characterization of Ultra-thin Metal Films for Nanofabrication Applications Matteo Todeschini*1, Henri Jansen1, Jakob B. Wagner1, Anpan Han1 and Alice Bastos Fanta1 1

DTU Danchip/Cen, Technical University of Denmark, Fysikvej, 2800 Kgs. Lyngby *E-mail: mattod@dtu.dk

Keywords: transmission Kikuchi diffraction, high T in-situ analysis, ultra-thin metal films.

The fabrication process of increasingly complex multi-material and multi-layer structures and devices in nanofabrication includes thermal treatments that might affect the nanostructure and stability of the films heavily. An important temperature-related problem for the application of such systems is solid-state dewetting1, the agglomeration of the film to form islands or nanoparticles when heated to sufficiently high temperatures, which can have a harmful effect on the performances and time-stability of the fabricated devices.

The fundamental insight in the driving forces for changes in crystal structure, grain size and crystallographic texture of such systems due to thermal treatments requires the introduction of innovative techniques having nanoscale resolution and being able to operate in a wide range of temperatures.

In this study, we demonstrate the in-situ capabilities of transmission Kikuchi diffraction (TKD) for the analysis of ultra-thin Au films at high temperature. The dewetting of an Au thin film into Au nanoparticles upon heating is followed with orientation mapping in a temperature range between 20°C and 900°C. The evolution of grain size and film texture and the growth of holes in the film are tracked throughout the process with high resolution, accuracy and statistical significance. Several sources of influence on the quality and resolution of the acquired TKD maps are investigated: disturbance from infrared radiation, maximum measurable Au thickness, loss of crystalline order, thermal drift during heating, plasma cleaning of the sample and thickness variation of the film.

A quasi in-situ TKD mapping is also used to observe the positive impact on the stability of the Au nanostructure at elevated temperatures due to the presence of Ti and Cr transition metals used as adhesion layers. The results show that the continuity of the Au film is preserved up to 500°C using either of the adhesion layers (Fig. 1), but also how Cr and Ti have a different impact on the final Au film nanostructure.

Figure 1: Inverse pole figure (IPF) quasi in-situ TKD maps showing the evolution of Au nanostructure without (top) and with Cr adhesion layer (bottom). The presence of the adhesion layer maintains the Au film continuous up to 500°C. [1] C.V. Thompson, Annu. Rev. Mater. Res. 42, 399–434 (2012).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark C-O5

Session: Materials Science and Energy Materials Substitution of Inherent Surface Terminations of 2D Titanium Carbide I. Persson*1, L-Å. Näslund1, J. Halim1, J. Palisaitis1, V. Darakchieva1, J. Rosén1, T. W. Hansen2, J. B. Wagner2, and P.O.Å. Persson1.

1

Department of Physics, Chemistry and Biology, Linköping University, SE-581 83 Linköping, Sweden 2 Center for Electron Nanoscopy, Technical University of Denmark, DK-2800 Lyngby, Denmark *E-mail: ingemar.persson@liu.se Keywords: in situ, ETEM, 2D materials, MXene, surface termination.

Two-dimensional (2D) materials exhibit an exceptionally high surface area per volume leading to an equally staggering number of available sites for surface reactions. MXenes (M= transition metal, X= C and/ or N) constitute a large family of 2D materials that possess a mixture of metallic and ceramic properties. These materials have attracted significant attention primarily because of their electrochemical properties, [1-2] photocatalysis, [3] and electromagnetic interference shielding (EMI) applications among others. [4] MXenes are derived from a 3D nanolaminated parent MAX phase (e.g. Ti3AlC2). In the MAX phase, a single atom thick A layer (e.g. Al) separates sheets of MX (e.g. Ti3C2). Chemical etching removes the separation layer and 2D MXene sheets are formed. However, the emerging MXene is subject to the adsorption terminating elements (Tx) originating from the etchant. [5] Presently, no means of substituting the inherent surface termination exist other than modification of the O, OH and F contents. [6] The ability to control the surface chemistry is critical for MXene property engineering.

In this contribution, we have investigated the limits for modification and tailoring of the surface functionalization of Ti3C2. This was conducted by exposing single or few layer Ti3C2Tx to a range of environments in an aberration corrected environmental transmission electron microscopy (ETEM). The evolution of Ti3C2Tx surfaces was followed by employing highresolution ETEM imaging, electron diffraction, electron energy-loss spectroscopy and residual gas analysis.

1 2 3

4 5

6

Lukatskaya, M. R. et al. Cation Intercalation and High Volumetric Capacitance of TwoDimensional Titanium Carbide. Science 341, 1502 (2013).

Naguib, M. et al. Two-Dimensional Nanocrystals Produced by Exfoliation of Ti 3 AlC 2. Adv. Mater. 23, 4248 (2011).

Yuan, W. et al. 2D-Layered Carbon/TiO2 Hybrids Derived from Ti3C2MXenes for Photocatalytic Hydrogen Evolution under Visible Light Irradiation. Advanced Materials Interfaces 4, 1700577-n/a, doi:10.1002/admi.201700577 (2017).

Shahzad, F. et al. Electromagnetic interference shielding with 2D transition metal carbides (MXenes). Science 353, 1137 (2016).

Dall'Agnese, Y. et al. High capacitance of surface-modified 2D titanium carbide in acidic electrolyte. Electrochemistry Communications 48, 118-122, doi:10.1016/j.elecom.2014.09.002 (2014).

Persson, I. et al. On the organization and thermal behavior of functional groups on Ti 3 C 2 MXene surfaces in vacuum. 2D Materials 5, 015002 (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark C-O6

Session: In Situ Nanoscale Microscopy of Processes In-situ visualizing the dynamic behaviors of nanocatalysts under gas environment 1

Yong Wang*1 School of Materials Science and Engineering, Zhejiang University, Hangzhou 310027 *E-mail: yongwang@zju.edu.cn Keywords: ETEM, catalysts, gas cell.

The environment, such as temperature, gas and pressure, has a significant impact on the catalyst surfaces, which may result in structural and electronic changes including gas adsorption, electron transfer and stress variation, making it challenging to accurately interpret the physical and chemical properties of nanocrystal catalysts. To address such an important issue, we introduce the environment into transmission electron microscope, which is supposed to work in high vacuum, to study the structural evolution and its impact on the physicochemical properties of nanocatalysts at the atomic scale under gas environment. In this talk, I will present our recent in-situ studies on nanocrystal catalysts under high temperature and/or gas environment (up to 105 Pa using gas-cell system) in TEM. The effect of the environment on nanocrystal catalysts will be discussed.

[1] Y. Jiang, H. Li, Z. Wu, Y. Wang, C. Sun and Z. Zhang, Angew Chem. Int. Ed. 55, 1242712430 (2016).

[2] X. Zhang, J. Meng, B. Zhu, J. Yu, S. Zou, Z. Zhang, Y. Gao, Y, Wang, Chem. Commun.

53, 13213-13216 (2017).

[3] W. Yuan, Y. Wang, H. Li, H. Wu, Z. Zhang, A. Selloni and C. Sun, Nano Lett. 16, 132137 (2016).

[4] Y. Jiang, Z. Zhang, W. Yuan, X. Zhang, Y. Wang, Z. Zhang, Nano Res. 11, 42â&#x20AC;&#x201C;67 (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-O1

Session: In Situ Nanoscale Microscopy of Processes In-situ Compositional Analysis of Catalyst Particle During GaAs Nanowire Growth in an Environmental TEM Carina B. Maliakkal*1,2, Daniel Jacobsson1,3, Marcus Tornberg1,2, Axel R. Persson1,3, Jonas Johansson1,2, Reine Wallenberg1,3 and Kimberly A. Dick1,2,3.

1

NanoLund, Lund University, Box 118, 22100, Lund, Sweden.

Solid State Physics, Lund University, Box 118, 22100, Lund, Sweden.

3 nCHREM and CAS, Lund University, Box 124, 22100, Lund, Sweden. *E-mail: carina_babu.maliakkal@ftf.lth.se 2

Keywords: nanowire, XEDS, catalyst composition.

III-V semiconductor nanowires (NWs) have potential widespread technological applications and their growth mechanisms are usually studied by post-growth analysis.[1] In the past few years, NWs have also been grown in environmental transmission electron microscopes (ETEMs) where their morphology and the kinetic processes have been monitored in-situ as they grow.[2-3] We advance this field further by studying the composition of the catalyst particles in an ETEM during the growth of GaAs NWs. We grow GaAs nanowires in a Hitachi HF3300S ETEM with B-COR-aberrationcorrector, and connected to a CVD system. A Si/Si3N4 MEMS heating chip is mounted on a single tilt holder that has two separate microtubes running to the holder tip for supplying gases. We use trimethylgallium and arsine as the Ga and As precursors respectively. Metallic nanoparticles which can alloy with at least one of the components of the NW and can act as catalysts are often used during NW growth. We use Au nanoparticles as it alloys with Ga. Chemical analysis of the Au-Ga particles is done by X-ray energy dispersive spectroscopy.

We studied the elemental composition of the catalyst as a function of the growth temperature and the ratio of the two precursor gases. On increasing temperature from 420 oC to 500 oC, at fixed precursor fluxes, the Ga content in the catalyst increases from about 23% to 35%. When the Ga precursor flow is increased, while maintaining a constant growth temperature and As precursor flow, again the Ga content increases in the catalyst (Fig. 1b).

High resolution videos recorded with high-frame-rates (20 fps) enables us to find growth rate for each bilayer, incubation times between layers etc. and to correlate it with the seed particle composition. Some representative videos will be presented. (a)

(b)

5nm Figure 1: (a) NW-catalyst interface (b) Catalyst composition at different Ga precursor flows.

[1] F. Glas, J.C. Harmand and G. Patriarche, Physical Review Letters 99, 146101 (2007).

[2] F. Lenrick, M. Ek, K. Deppert, L. Samuelson, and L.R. Wallenberg, Nano Research 7, 1188-1194 (2014).

[3] D. Jacobsson, F. Panciera, J. Tersoff, M.C. Reuter, S. Lehmann, S. Hofmann, K.A. Dick and F.M. Ross e, Nature 531, 317-322 (2016).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-O2

Session: In Situ Nanoscale Microscopy of Processes

Observing Catalyst Structures and Dynamics at Atomic-Resolution M. Ek1, L. P. Hansen1, F.-R. Chen2, D. van Dyck3, C. Kisielowski4, J. R. Jinschek5, S. Helveg*1.

1.

Haldor Topsoe A/S, Haldor Topsøes Allé 1, DK-2800 Kgs. Lyngby, Denmark.

Department of Engineering and System Science, National Tsing-Hua University, Hsin Chu, Taiwan.

3.

Departments of Physics, EMAT, University of Antwerp, 2020 Antwerp, Belgium.

4.

Molecular Foundry, Lawrence Berkeley National Laboratory, Berkeley CA 94720, USA. 5.

Department of Materials Science and Engineering & Center for Electron Microscopy and Analysis (CEMAS), Ohio State University, Ohio 43212, USA. *E-mail: sth@topsoe.com 2.

Keywords: Heterogeneous catalysts, in situ studies, transmission electron microscopy.

Over the past decade, electron microscopy has become indispensable for studying heterogeneous catalysts. The ability to acquire atomic-resolution images with single-atom sensitivity has opened up for unprecedented insight into catalyst structures and dynamics.

However, the atomic-level observations require intense electron illumination that generally alters the catalysts [1-3]. Electron-induced alterations are particular pronounced at the catalyst surface as they expose a variety of sites of reduced atomic coordination. In the quest to suppress electron-induced alterations and to enable chemical meaningful observations, it therefore becomes mandatory to exercise control over the electron dose, dose-rate and energy.

Here, we demonstrate low dose-rate in-line electron holography as a viable concept for atomic-resolution observations of catalysts in a genuine state [2-5]. The imaging scheme employs bright field transmission electron microscopy (TEM) to efficiently detect single atoms using the fewest elastically scattered electrons. The image acquisition is done with low electron dose-rates of down to 1-100 e-Å-2s-1 to induce weak object excitations and provide time for reversible object restoration between successively delivered electrons. As a result, the individual images are noise-dominated, so the acquisition of focal image series for in-line holography becomes particular attractive to correct residual aberrations and recover the exit wave (EW) function with enhanced signal.

The significance of low dose-rate in-line electron holography is a suppression of beaminduced sample alterations that enables a quantitative interpretation of the EW. The low doserate in-line holography concept will be illustrated by observations of a Co3O4 photo-oxidation catalyst and a Co-Mo-S hydrotreating catalyst, and even by in situ observations of a VOx/TiO2 NOx removal catalyst under reactive gas conditions [4-7].

[1] S. Helveg, J. Catal. 328, 102 (2015).

[2] C Kisielowski et al, Phys Rev B 88, 024305 (2013).

[3] S. Helveg et al, Micron 68, 176 (2014).

[4] C. Kisielowski et al, Adv. Struct. Chem. Imag. 2, 13 (2016).

[5] F.-R. Chen, D. van Dyck, C. Kisielowski, Nature Comm. 7, 10603 (2016).

[6] F.-R. Chen et al (2018) in prep.

[7] M. Ek et al, Nature Comm. 8, 305 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-O3

Session: In Situ Nanoscale Microscopy of Processes In Situ Gold Nanoparticle Formation in a Free-Standing Ionic Liquid Layer Triggered by Heat and Electron Irradiation Debora Keller*1, Trond Henninen1, and Rolf Erni1.

1

Electron Microscopy Center, Empa, Swiss Federal Laboratories for Materials Science and Technology, Überlandstrasse 129, CH-8600 Dübendorf, Switzerland. *E-mail: debora.keller@empa.ch Keywords: ionic liquid, in situ, heating, gold, nanoparticle growth.

Ionic liquids are known to withstand high vacuum conditions while being in their liquid state.

This peculiarity makes them very promising for use as a liquid medium to study nanoparticle (NP) nucleation and growth in situ at high spatial resolution in transmission electron microscopy (TEM). So far, a few studies demonstrated NP nucleation and growth by electron irradiation in ionic liquids [1,2], but still only very little information about such complex systems’ behavior in the TEM is available.

In this work we study the formation of gold NPs in a free-standing ionic liquid layer, triggered by heat and electron irradiation. A sample containing NaAuCl4 and 1-butyl-3methylimidazolium chloride ([Bmim][Cl]) is investigated with a Fusion holder from PROTOCHIPS which allows for heating in situ in the TEM. At room temperature, where [Bmim][Cl] is still in a solid state, no particle growth is observed in scanning (S-)TEM mode, independent from the irradiation conditions. However, Au particles form slowly when the sample is heated well above its melting temperature of 80°C. The growth of the Au particles is accelerated as the temperature increases and two different morphologies are typically observed: firstly, small spherical particles precipitate and secondly, larger, facetted particles are built such as decahedra, cubes or tetrahedra. However, the particle growth mechanism and the ultimate particle size and shape are not only determined by temperature but also by other factors, such as the electron dose (total dose and dose rate), time and the liquid layer’s geometry. In order to characterize the growth process in more details and to distinguish the influence of different factors, further studies are still necessary. Moreover, capturing the NP growth process in detail could provide valuable information in future about the evolution of the different particles and the possible transformations from one particle morphology into another.

Figure 1: (a) Experimental setup, (b) Au NP grown in situ in ionic liquid.

[1] Uematsu et al., J Am Chem Soc 136, 13789–13797 (2014).

[2] Kimura et al., J Am Chem Soc 136, 1762–1765 (2014).

[3] This project has received funding from the European Research Council (ERC) under EU’s Horizon 2020 research and innovation program (Grant Agreement No. 681312).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-O4

Session: In Situ Nanoscale Microscopy of Processes Atomic-resolution imaging of heterogeneous catalysts at work S. Helveg*1, C. F. Elkjær1, S. P. Jespersen1, S. Vendelbo1, B. Hendriksen2, L. Mele2, P. Dona2, J. F. Creemer3, P. Kooyman4, I. Chorkendorff5, C. Damsgaard5, J. R. Jinschek6 1.

Haldor Topsoe A/S, Haldor Topsøes Allé 1, DK-2800 Kgs. Lyngby, Denmark.

Thermo Fisher Scientific, Achtseweg Noord 5, 5651GG Eindhoven, The Netherlands.

3.

DIMES-ECTM, Delft University of Technology, 2600 GB Delft, The Netherlands.

4.

Department of Chemical Engineering, University of Cape Town, South Africa.

5.

Department of Physics, Technical University of Denmark, Denmark.

6.

Department of Materials Science and Engineering, Ohio State University, USA. . *E-mail: sth@topsoe.com 2.

Keywords: Heterogeneous catalysis, nanoreactor, TEM, operando, in situ.

The ability to acquire atomic-resolution images of catalysts opens up for unprecedented new information about the catalyst surface that benefits the understanding of structure-sensitivity in catalysis [1]. However, surfaces of most materials tend to restructure in reactive gas environments and such changes have a profound feedback on catalysts’ properties. It has therefore remained a key milestone in catalysis and surface sciences to functionalize electron microscopy for operando studies in which the catalyst surface structure and properties are simultaneously evaluated at the atomic-scale and under relevant reaction conditions.

Here, we report on a nanoreactor system that enables combined electron microscopy and functional measurements of catalysts [2-6] and is available from Thermo Fisher Scientific.

The nanoreactor is a micro-electro-mechanical system device integrating a 5-µm-high, one-pass and bypass-free gas-flow channel, a microheater and an array of 15-nm-thick electrontransparent windows of silicon nitride (Figure 1) [4]. This nanoreactor is operational with variable gas flows (0-0.1 sccm/min), ambient pressure levels (> 1 bar) and elevated temperatures (25-700 oC), while maintaining the inherent 1Å resolution in modern electron microscopes. Moreover, mass spectrometry of gas exiting the reaction zone is made possible by the small reaction volume of ca. 0.3 nL and reaction calorimetry is accessed from the power compensation used for isothermal operation of the microheater [6].

The nanoreactor performance and application will be showcased by observations of Pt nanoparticles under oxidizing and reducing reaction conditions. Specifically, high-resolution electron microscopy images were acquired of the Pt nanoparticles under exposure to reactive gas environments synchronously with mass spectrometry and calorimetry data. Based on such observations, structure-activity relationship will be outlined for e.g. the steady-state and oscillatory catalytic oxidation of carbon monoxide over Pt nanoparticles [6,7]. These examples demonstrate exciting possibilities for including information about the exposed surface sites into the description of dynamic properties and functions of catalysts.

[1] S. Helveg, J. Catal. 328 (2015), p. 102.

[2] J.F. Creemer et al, Ultramicroscopy 108 (2008), p 993.

[3] J.F. Creemer et al, J. Microelectromech. Syst. 19 (2010), p. 254.

[4] J.F. Creemer et al, Proceedings of MEMS 2011 (2011), p. 1103.

[5] S.B. Vendelbo et al, Ultramicroscopy 133 (2013), p. 72.

[6] S.B. Vendelbo et al, Nature Materials 13 (2014), p. 884.

[7] C.F. Elkjær et al, in preparation.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P1

Session: In Situ Nanoscale Microscopy of Processes 3D Chemical Analysis of Inorganic and Organic Nanostructures using ToF-SIMS and In-situ SPM Sven Kaysera, Ewald Niehuisa, Rudolf Moellers a, Felix Kollmer a, Derk Rading a, Henrik Arlinghaus a, a Andreas Duetting , Raphaelle Dianoux b, Adi Scheidemann b a b

ION-TOF GmbH, Muenster, Germany NanoScan AG, Duebendorf, Switzerland Email: ewald.niehuis@iontof.com

TOF-SIMS is known to be an extremely sensitive surface analysis technique which provides elemental as well as comprehensive molecular information on any kinds of solid surfaces. In combination with conventional low energy oxygen or cesium sputtering, 3D structures can be analysed with a lateral resolution of down to 50 nm and a depth resolution in the nm range.

With the advent of large gas cluster ion beams (GCIB) [1], the 3D capability of the TOF-SIMS was extended to complex organic materials and devices [2]. Inherent to all 3D SIMS data is a z-axis with a native time scale instead of a length scale. A starting topography of the initial sample surface as well as an evolving topography due to different sputter rates of the compounds cannot be identified by the technique and lead to major distortions of 3D data sets.

The sputter rates of the various inorganic and organic materials are very different in particular for large gas cluster sputtering [3] and can be strongly influenced by radiation damage of organic materials [4].

Scanning Probe Microscopy (SPM) provides the required complementary information on the surface topography down to the nanometer level. Beyond that, SPM can provide valuable information on physical properties if the cantilever is operated in the appropriate dynamic operation modes.

We integrated an SPM unit into a ToF-SIMS instrument in some distance from the SIMS analysis position. The core piece of the new instrument is a piezo driven stage which moves the sample between the TOF-SIMS and the SPM analysis position with high precision and speed.

The SPM unit with a beam deflection design is mounted on a 3-axis linearized scanner with a scan range of 80 x 80 x 10 µm³. This flexure stage scanner has a very small out-of-plane motion and yields very accurate information on the surface topography. The SPM is also required to measure the sputter crater depth with high precision. For crater sizes of several hundred µm, a special long distance surface profiler mode was developed to measure the correct shape and depth of the sputter crater. In order to measure a variety of physical sample properties, the SPM can be operated in dynamic modes including KPFM, conductive AFM and MFM. In this paper we will present various examples highlighting the strength of this novel combined instrument and its potential for a wide range of applications. The examples include fundamental studies on the sputtering of organic and hybrid materials with various sputter beams like Ar, O2 and SF6 gas clusters as well as applications for the characterization of 3D objects like OLEDs and biological single cells.

[1] S. Ninomiya, K. Ichiki, H. Yamada, Y. Nakata, T. Seki, T. Aoki and J. Matsuo, Rapid Commun. Mass Spectrom. 23, 3264 (2009)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P2

Session: In Situ Nanoscale Microscopy of Processes [2] E. Niehuis, R. Moellers, D. Rading, H.-G. Cramer, R. Kersting, Surf. Interface Anal. 45, (2013) 158 [3] M.P. Seah, J. Phys. Chem. C, 2013, 117(24), pp 12622-12632 [4] T. Conard, A. Franquet, D. Tsvetanova, T. Mouhib, W. Vandervorst, Surf. Interface Anal. 45, (2013) 406

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P2

Session: In Situ Nanoscale Microscopy of Processes Direct in Situ Monitoring of Nanoalloy Transformation Pathways Min Tang*1, Beien Zhu2, and Yong Wang1.

1

2

State Key Laboratory of Silicon Materials and Center of Electron Microscopy, School of Materials Science and Engineering, Zhejiang University, Hangzhou 310027 (China).

Division of Interfacial Water and Key Laboratory of Interfacial, Physics and Technology, Shanghai Institute of Applied Physics, Chinese Academy of Science, Shanghai 201800 (China). *E-mail: mintang@zju.edu.cn

Keywords: transformation pathways, Pd-Pt core-shell nanoparticles, in-situ STEM, alloying.

Nanoalloys have attracted lots of attention for their wide applications in materials, optics, catalysis and biomedicines, which largely rely on their composition-, size-, and shapedependent properties [1]. Since these properties change dynamically with environment [2], the knowledge of the complex transformation pathways of individual nanoalloys is highly demanded. In this work, in order to monitor the whole transformation trajectory of the bimetallic nanoalloy (Pd-Pt), we employed the in-situ aberration-corrected scanning transmission electron microscopy observations at atomic resolution in a long time annealing treatment. We found the transformation from core-shell to solid-solution structure is a multistep and temperature-dependent pathway, which includes monometallic-surface refacetting, bimetallic-surface refacetting, and alloy mixing. The mechanism is further rationalized by density functional theory and multi-scale modeling. This study offers a fundamental insight of the structure evolution of nanoalloys, which is beneficial for the development of the functionalized nanoparticles with kinetic stability.

Figure 1: (a-f) Atomic-resolution HAADF-STEM images of a single Pd-Pt NP acquired at different temperature and annealing times. (g-l; m-r) Corresponding details of {110} and {100} facets, respectively.

[1] R. Ferrando and J. Jellinek, Chem. Rev. 108, 845â&#x20AC;&#x201C;910 (2008).

[2] Y. Jiang, H. Li, and Y. Wang, Angew. Chem.Int. Ed. 55, 12427-12430 (2016).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P3

Session: (In Situ Nanoscale Microscopy of Processes)

Temperature Dependent Quasimolten Crystallinity of sub-nm Pt and Au Clusters Observed in 3D by Fast Dynamic STEM Trond Henninen*1, Marta Bon1, Daniele Passerone1, Rolf Erni1 1

Empa, Swiss Federal Laboratories for Materials Science and Technology, CH-8600 Dübendorf, Switzerland *Email: trond.henninen@empa.ch

Keywords: Quantitative STEM, sub-nm, quasimolten, atomic clusters, in-situ heating The formation of solid matter is typically modelled by classical nucleation theory, assuming the solid forms a spherical nucleus. However, during the initial phase at sub-nm scale, non-spherical atomic clusters form, with atomic structures deviating from bulk crystals.The aim of this work is to study the metastable structures and atomic dynamics of sub-nm clusters, as individual atoms combine to form stable nm-sized clusters.

Plasma sputtered samples of Pt or Au with a homogeneous coverage of single atoms and clusters up to ~2nm, were studied using a probe corrected FEI Titan Themis at 300 kV. The microscope was optimized for fast dynamic STEM imaging and low radiation dose, recording at ca 2.5, 15 and 150 fps with doses ranging correspondingly from 5*103 e/Å2s to 2*106 e/Å2s. Heating experiments were done from room temperature to 500°C. 3D reconstruction was done by building atomic models assisted by StatSTEM[1] atom counting based on images from multiple orientations, recorded as the clusters rotate.

At room temperature, the clusters have a largely flat, amorphous structure, 1-2 atoms tall, as if the clusters wet the carbon surface in a droplet-like manner. Counterintuitively, the clusters crystallize when heated up; forming close-packed quasi-molten crystals with a more spherical shape, almost as tall as wide. This crystallization is reversible upon cooling the clusters to room temperature, showing that the amorphous and crystalline phases are equilibrium phases at their respective temperatures. We directly observed multiple different structures forming from the same specific atomicity clusters. For example, for 13-atom Pt clusters, four different identified structures are shown in the figure. Notably, the cube-like fcc-type structure was abnormally stable during e-beam irradiation compared to the others. A larger 18-atom cluster of the same cubic type was also abnormally stable. This type of cubic clusters has been simulated to be the favoured growth pathway for Pt clusters[2]; however, this is in contrast to the majority of theoretical work which agree that spherical closed shell icosahedral-type clusters are the most stable.

Figure: Denoised STEM images (top) and atomic models (bottom) of four different crystalline structures for Pt13 clusters. a,e cube-like fcc. b,f fcc cuboctahedron. c,g hcp. d,h icosahedron [1] A. De Backer et al., Ultramicroscopy 171, (2016) [2] A. Nie et al., Int J Quantum Chem 107, (2007) Project funding: ERC under EU’s Horizon 2020 programme (grant No. 681312)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P4

Session: In Situ Nanoscale Microscopy of Processes Formation of Prussian Blue Nanocubes Studied by Liquid Cell Transmission Electron Microscopy Hongyu Sun*1, Murat Nulati Yesibolati1, Minwei Zhang2, and Kristian Mølhave1.

1

DTU Nanotech, Technical University of Denmark, Kgs. Lyngby, Denmark.

2 DTU Chem, Technical University of Denmark, Kgs. Lyngby, Denmark. *E-mail: hsun@nanotech.dtu.dk

Keywords: solution synthesis, prussian blue, liquid cell transmission electron microscopy.

Metal Organic Frameworks (MOFs) are materials with extreme porosity and are usually synthesized in solution by joining metal ions with organic linkers [1]. As typical MOFs, prussian blue (PB) and analogues have been explored for applications in sensors, energy conversion and storage. PB materials often fail to realize their expected performance, possibly due to poor understanding and control of the PB crystal growth [2]. Recently, liquid cell transmission electron microscopy (LCTEM) has been successfully applied to study a wide range of processes in solution with subnanometer scale spatial resolution [3-5]. Compared to ex-situ characterization methods, LCTEM provides direct evidence about crystal evolution in liquid phase through real-time observation.

In this work, we employ in-situ flow LCTEM for real time observation of the evolution of PB nanocube synthesis in solution. By using continuous and successive flows of reagent solutions (first HCl, 30 uL/min and then H2O2 solution, 50 uL/min) into the liquid cell containing K4[Fe(CN)6], we observed the formation of dispersed PB nanocubes with typical size ranging from 20 to 70 nm (Fig. 1a). Sometimes, the PB nanocubes assembled into chainlike morphology (see the white arrows in Fig. 1b). Although we carefully controlled the electron flux (~ 1.59 e/Å2s) to reduce the beam effect during imaging, the PB nanocubes were prone to completely dissolving in the solution (Fig. 1c) within one minute of irradiation.

Under the same imaging conditions, dry PB nanocubes only transformed from the initial crystalline into amorphous structure (Fig. 1d). The results demonstrate LCTEM method is useful to study the PB synthesis in solution, making it possible to provide insight to obtain better control of the crystalline of PB materials towards advanced applications.

Figure 1: (a, b) Typical TEM images of PB nanocubes in liquid cell, (c) Time series of TEM images of PB nanocubes in liquid (incident electron flux: 1.59 e/Å2s), (d) TEM images of dry PB nanocubes before and after 1 min beam irradiation.

[1] N. Stock, and Shyam Biswas, Chem. Rev. 112, 933–969 (2012).

[2] Y. H. Lu, et al, Chem. Commun. 48, 6544–6546 (2012).

[3] F. M. Ross, Science 350, aaa9886 (2015).

[4] J. P. Patterson, et al, J. Am. Chem. Soc. 137, 7322–7328 (2015).

[5] S. A. Canepa, et al, J. Phys. Chem. C 122, 2350–2357 (2018).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P5

Session: In Situ Nanoscale Microscopy of Processes Electrochemical impedance spectroscopy TEM implementation for a model solid oxide electrolysis cell Søren B. Simonsen*1, Fabrizio Gaulandris1, Jakob B. Wagner2, Kristian Mølhave 3, S. Sanna1, Shun Muto4, Luise Theil Kuhn1.

1

Department of Energy Conversion and Storage, Technical University of Denmark, DK-4000 Roskilde, Denmark, 2 DTU Danchip/Cen, Center for Electron Nanoscopy, Technical University of Denmark, DK2800 Kgs. Lyngby, Denmark, 3 Department of Micro and Nanotechnology, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark, 4 Institute of Materials and Systems for Sustainability, Nagoya University, Nagoya, Aichi, Japan *E-mail: sobrs@dtu.dk Keywords: in situ TEM, electrochemical impedance spectroscopy, solid oxide electrolysis cells, solid oxide fuel cells.

The solid oxide electrolysis cell (SOEC) is a promising technology for conversion of electrical energy to chemical energy (hydrogen or hydrocarbons) for storage [1]. The operation conditions (≥ 800°C in reactive gasses) induce nanoscale changes in the active materials thereby changing the overall cell performance. Post mortem analysis is insufficient to understand the time, temperature and electrical potential dependencies of these changes. It is, however, a great challenge to conduct in situ electron microscopy on SOECs since this requires: 1) that the hard and brittle ceramic cells are thinned to electron transparency, 2) that the cells are carefully designed to allow for characterization of the layer interfaces, 3) and that the cells are characterized during exposure of reactive gasses, 4) electrical potentials and 5) high temperatures.

Here we present a TEM/STEM study where such in situ experiments were performed on symmetric model SOECs composed of materials commonly used in state-of-the-art SOECs, i.e. La0.6Sr0.4CoO3-δ (LSC) electrodes and a Zr0.8Y0.2O2-δ (YSZ) electrolyte (fig. 1). This was achieved in an environmental TEM in combination with both custom-made and commercial heating/biasing TEM holders. In this work we extended the TEM in situ tool box by implementing electrochemical impedance spectroscopy (EIS) on the model SOECs in the TEM. Preliminary EIS-TEM experiments show a decrease in resistance over the model cells. This trend is expected since the oxygen ion conductivity increases with temperature and similar trends are observed for full scale SOECs.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P6

Session: In Situ Nanoscale Microscopy of Processes

Figure 1: TEM image of the model SOEC with drawings added to illustrate the in situ conditions: gas flow, elevated temperature and electrical potential.

Acknowledgment The Danish Council for Independent Research is acknowledged for providing funding for the project In situ transmission electron microscopy on operating electrochemical cells – TEMOC grant no. DFF – 4005-00247.

[1] [2] [3]

N.Q. Mitili, M.B. Mogensen, Interface. 22 55-62 (2013).

F. Gualandris, S.B, Simonsen, J.B. Wagner, S. Sanna, S. Muto, L.T. Kuhn, ECS Transactions 75 123-133 (2017).

M.J. Jørgensen, S. Primdahl, M.B. Mogensen, Electrochimica Acta 44 4195-4201 (1999).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P6

Session: In Situ Nanoscale Microscopy of Processes New Insights in CMOS Based TEM Detector Reza Ghadimi*1, Marco Oster1, Dominic Tietz1, Andreas Wisnet1, and Hans Tietz1.

1

TVIPS GmbH, Eremitenweg 1, 82131 Gauting, Germany. *E-mail: reza.ghadimi@tvips.com

Keywords: TEM Camera, In-situ, Precession Diffraction Tomography, EELS. The TemCam-XF416 is the latest development of TVIPS featuring an entirely new 4k×4k sensor design, which simultaneously satisfies different criteria: Large pixel size (16 µm x 16 µm), single electron sensitivity (high signal-to-noise ratio), fast acquisition, extended dynamic range, high spatial resolution and applicability for a wide range of acceleration energies (8 kV-400 kV).

The new pixel design offers a physical dual readout mode, which combines single electron sensitivity and ultra-high dynamic, revealing new insights in nano-scaled materials by cryo electron diffraction tomography on crystals (MicroED).

The high frame rate (50 fps full-frame, up to 400 fps subarea) combined with on-chip correlated double sampling allows dose fractioning to reduce drift while simultaneously offering single electron counting to reduce the Landau noise (Figure 1). Furthermore, the TemCam-XF416 can be utilized as an ideal detector for in-situ experiments, e.g. in combination with holders by DENSsolutions.

Experimental results demonstrate the ability of the TemCam-XF416 to be used in a wide range of electron energies - even at 20 kV single electrons can be observed.

Figure 1: (left) Single electron events at 20 kV, as recorded by a TemCam-XF416. (Right) reconstructed image by electron counting (total dose: 4 ē/Å2).

Figure 2: In-situ heating experiment on Au nanoparticles by DENSsolution heating holder.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P7

Session: In Situ Nanoscale Microscopy of Processes Dynamics of Nanostructures: A quantitative approach using in situ Electron Microscopy Thomas W. Hansen*1, Pei Liu1, Jacob Madsen2, Monia Runge Nielsen1, Jakob Schiøtz2, Jakob B. Wagner1.

1

2

Center for Electron Nanoscopy, Technical University of Denmark, DK-2800, Kgs. Lyngby, Denmark Department of Physics, Technical University of Denmark, DK-2800, Kgs. Lyngby, Denmark *E-mail: twh@cen.dtu.dk Keywords: nano particles, in situ TEM, quantitative analysis, electron beam effects

Surfaces of nanoparticles represents a topic of interest in several fields. For example, the activity of heterogeneous catalysts strongly depends on the structure of these surfaces. The nature of these surfaces is a function of the environment to which the nanoparticle is exposed, i.e. surrounding gases and temperature. Detailed quantitative information of the surfaces under operating conditions is inherently difficult to access due to limitations of characterization equipment. Advances in electron microscopy such as MEMS heaters, reaction cells and fast cameras have provided new possibilities for in situ characterization at the atomic scale [1] giving access to even more detailed information. Knowledge of the imaging conditions is a prerequisite in order to extract meaningful results. Application of automated analysis can provide strong and robust characterization of nanoparticle structure and dynamics can be achieved. However, automated image analysis does have its pitfalls.

Whereas this is also true for manual analysis, at least the human factor can be ruled out.

In the present study, we investigate the surface structure and dynamics of gold nanoparticles under varying atmospheres. Using environmental high-resolution transmission electron microscopy, gold nanoparticles supported on cerium dioxide have been imaged under varying conditions.

Firstly, the equilibrium shape under oxidizing and reducing conditions was investigated.

Under vacuum, the surfaces appeared stable meaning that no change in column occupancy, see Figure 1. The figure shows both the occupancy degree of each column as well as the diffusion frequency. As the samples are exposed to oxygen at 4.5 Pa, the atomic columns on both {111} and {001} facets start diffusing. When the samples are exposed to hydrogen, only the atomic columns on the {001} facets diffuse. These observations indicate that as expected, different gas molecules interact differently with different facets. A possible explanation of the observed events is the higher desorption temperature of oxygen (> 415 K) compared to hydrogen (~120 K).

Under exposure to 4.5 Pa carbon monoxide and 300°C, surface layers start shifting in a concerted fashion, and a crystal twin parallel to the nanoparticle/support interface forms and moves dynamically resulting in at least two distinct configurations of the nanoparticle. In each configuration, the surface strain on the nanoparticle was measured using a newly developed routine, see Fig. 2 [2]. In the untwinned configuration (upper pane), the lower (111) facet shows strong outwards relaxation. In the twinned configuration (lower pane), the corner sites show an outward relaxation. These observations strongly suggest that the catalytically active sites are dynamic rather than static entities.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P8

Session: In Situ Nanoscale Microscopy of Processes

Figure 1: Quantification of occupancy and hopping frequency on the surface of a gold nanoparticle at room temperature.

Figure 2: Surface strain on a gold nanoparticle in CO at 300°C. [1] T.W. Hansen and J.B. Wagner, eds. “Controlled Atmosphere Transmission Electron Microscopy”. 1 ed. 2016, Springer.

[2] J. Madsen, et al, Advanced Structural and Chemical Imaging 3, 14 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P8

Session: In Situ Electron Microscopy Imaging nanoparticle formation in heterogeneous catalysts Christian F. Elkjær*1, Roy van den Berg1, Jens Sehested1 and Stig Helveg1.

1

Haldor Topsoe A/S, Haldor Topsøes Allé 1, DK-2800 Kgs. Lyngby, Denmark *E-mail: chre@topsoe.dk

Keywords: TEM, in situ, nanoparticles, reduction, beam-sample interactions, catalysis.

In heterogeneous catalysis, chemical conversions typically proceed on the surface of nanometer-sized particles located on a porous support structure. The catalytic behavior depends on the size and spatial arrangement of these nanoparticles and the synthesis process is therefore of utmost importance for the final catalyst. A nanoparticle-support system may be formed from a homogeneous solid precursor where the nanoparticles nucleate and grow during a reduction process. Detailed insight into the growth of the nanoparticles is beneficially obtained using transmission electron microscopy (TEM) that enables individual nanoparticle to be tracked in situ. To ensure the chemical relevance of the observations, it is imperative to understand and minimize the influence of the electron beam on the process to obtain useful and quantitative data. Here we present a procedure that enable observations of thermally driven nanoparticle formation in beam-sensitive materials with negligible contribution from the electron beam.

The procedure will be illustrated by e.g. the reduction of Cu phyllosilicate[1]. Thermal and beam induced reduction are competing, so several strategies were used to first isolate and evaluate the effect of the electron beam. To do this, it was necessary to develop quantitative metrics to compare the evolution of different areas exposed with different dose rate or total dose. These metrics were the rate of particle growth and final particle size distribution (PSD).

Illumination prior to reduction e.g. had a significant influence on the final PSD. The strategy to minimize the effect of the electron illumination had several components: i) no illumination prior to reduction, ii) very low dose rates (1 e-/(Å2s)), iii) increased rate of thermal reduction and iv) comparison of multiple areas illuminated with different total dose. With this strategy, we acquired time-lapsed image series of individual nanoparticles with no detectable influence of the electron beam. An example of a time-lapsed image series, providing the location and size of individual nanoparticles is shown in Fig. 1. These image series combined with modeling showed that the reduction proceeded via an autocatalytic route.

Figure 1: Frames from time lapsed image series acquired during reduction in 1 mbar H2 at 280 °C. The imaging was performed according to a scheme that minimized the influence of the electron beam on the reduction process, thus providing quantitative data of the thermal reduction.

[1] Roy van den Berg, Christian F. Elkjær, Cedric J. Gommes, Ib Chorkendorff, Jens Sehested, Petra E. de Jongh, Krijn P. de Jong, and Stig Helveg, J. Am. Chem. Soc. 138 (2016)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P9

Session: In Situ Nanoscale Microscopy of Processes Ordered carbon van der Waals heterostructures Kimmo Mustonen*1, Rasim Mirzayev1, and Aqeel Hussain2, Christof Hofer1, Mohammad R. A. Monazam1, Esko I. Kauppinen2, Toma Susi1, Jani Kotakoski1 and Jannik C. Meyer1.

1

2

Faculty of Physics, University of Vienna, Boltzmanngasse 5, A-1090 Vienna, Austria.

Department of Applied Physics, Aalto University School of Science, P.O. Box 15100, FI00076 Aalto, Finland. *E-mail: kimmo.mustonen@univie.ac.at

Keywords: two-dimensional materials, van der waals heterostructures, single-walled carbon nanotubes, scanning transmission electron microscopy.

Two-dimensional (2D) materials have considerably expanded the field of materials science during the past decade. Even more recently, various 2D materials have been assembled into vertical van der Waals heterostructures (vdWHs), and it has been proposed to combine them with other low-dimensional structures to create new materials with hybridized properties. We have recently demonstrated self-ordered vdWHsâ&#x20AC;&#x2122; by suspending monolayer of C60 molecules in between two graphene sheets to create a buckyball sandwich structure [1], and by drydepositing [2,3] single-walled carbon nanotubes (SWCNTs) on monolayer graphene [4] excited to minimum energy orientation by laser irradiation in ultra-high vacuum [5]. We studied the structures in a scanning transmission electron microscope and observed, among other things, the diffusion of C60 molecules on the edges of hexagonally close-packed islands, self-ordering of SWCNTs on graphene and the emergence of one-dimensional grooves and radial deformation of carbon nanotubes as a result of the interfacial energy minimization in such systems. These materials may prove useful as in form of molecular reaction chambers and in creation of periodic potentials on 2D crystals.

[1] R. Mirzayev et al., Science Advances 3, 1700176 (2017) [2] K. Mustonen et al., Appl. Phys. Lett. 107, 013106 (2015) [3] P. Laiho et al., ACS Appl. Mater. Interfaces 9, 20738â&#x20AC;&#x201C;20747 (2017) [4] K. Mustonen et al., Graphene-carbon nanotube heterostructures: Self-alignment and deformations (submitted) [5] M. Tripathi et al., Rapid Research Letters 11, 1700124 (2017)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P10

Session: In Situ Nanoscale Microscopy of Processes Designing an Exchangeable Biprism for In-Situ Electron Holography Jes Ærøe Hyllested*1, Murat N. Yesibolati2, Flemming Jensen1, Jakob B. Wagner1 and Takeshi Kasama1 DTU Cen/Danchip, Technical University of Denmark, Kgs. Lyngby, Denmark 2 DTU Nanotech, Technical University of Denmark, Kgs. Lyngby, Denmark. *E-mail: Jehy@dtu.dk

1

Keywords: Biprism, MEMS, Electron holography, Environmental TEM, Instrumentation Off-axis electron holography is a TEM technique that allows for imaging magnetic and electric fields in materials quantitatively with nanometer spatial resolution. As such, it would be attractive to use electron holography in combination with environmental TEM (ETEM) with great potential for various applications: E.g., studies on changes in magnetic structures under reduction/oxidation conditions, charge distributions in catalytic nanoparticles under the presence of various gases, and electric field distributions in working fuel cells.

The conventional biprism is designed for high vacuum and ‘clean’ conditions in a TEM column. The pressure in ETEM conditions is higher by six orders of magnitude than that in a normal TEM and a significant number of gas molecules exist in the TEM column. Therefore, the conventional biprism device may not survive for a long period and may be contaminated by residue gas molecules. For these reasons, an easily exchangeable biprism device, with a high stability and resistance against corrosive gases would be required. Therefore, we have designed and fabricated an exchangeable biprism device using MEMS-based fabrication technology.

Figure 1 shows a schematic design of a MEMS-based biprism chip, where an Au/Si biprism wire with a vacuum space on both sides of the biprism wire will be fabricated. The optimal dimensions of the biprism wire and the vacuum space was calculated using COMSOL modelling software in order to generate uniform electric fields, where ’two' electron beams are deflected towards one another for making an interference overlap region. The biprism chip is mounted on a home-made biprism rod, which is inserted into the selected-area aperture port of a microscope column (Fig. 2). The rod also holds two spaces for mounting selected area apertures. This biprism device is made without a rotational functionality to achieve better mechanical stability, which would be desired for in-situ high-resolution electron holography of catalytic nanoparticles under ETEM conditions.

Vacuum

Biprism wire

100µm Ground electrodes

y x

Figure 1: Design of the electron biprism chip.

Figure 2: Schematic illustration of the electron biprism rod.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P11

Session: In Situ Nanoscale Microscopy of Processes Real time Observation of Dissolution and Regrowth Dynamics of MoO2 nanowires Wentao Yuan1, Ze Zhang1, Chenghua Sun2, and Yong Wang*1.

1

2

Center of Electron Microscopy and State Key Laboratory of Silicon Materials, School of Materials Science and Engineering, Zhejiang University, Hangzhou 310027, China.

Department of Chemistry and Biotechnology, Faculty of Science, Engineering & Technology, Swinburne University of Technology, Hawthorn, VIC, Australia. *E-mail: yongwang@zju.edu.cn Keywords: oxide surface, oscillatory behavior, dissolution kinetics, environmental transmission electron microscopy (ETEM)

Understanding anisotropic properties and stability of nanocrystals is of great importance to synthesize size- and morphology-controlled nanomaterials. Although great efforts have been devoted to revealing the fundamental mechanism of dissolution in solution or vacuum, gas involved dissolution, as well as regrowth dynamics of oxide nanocrystals at elevated temperatures, are rarely studied. Here, using an environmental transmission electron microscopy (ETEM), we performed an in situ study of the dissolution and regrowth dynamics of MoO2 nanowires under oxygen. Our in situ observation revealed an oscillatory dissolution of nanowire-tip facets and simultaneous layer-by-layer regrowth of sidewall facets, which result in a shorter and wider nanowire. Through density functional theory (DFT) calculations, we found oxygen-loss in the tip facets can be induced by electron beam irradiation, leading to change of the preferential growth facets and morphology reshaping of MoO2 nanowire.

Figure 1: (a) and (b) Sequential ETEM images and the corresponding models showing oscillatory morphology change of the (100) tip facet of a MoO2 nanowire. (Temperature: 600 o C; Oxygen pressure: 5Ă&#x2014;10-2 Pa; and continuous e-beam irradiation) (c) Plot of the dissolved area of (100) facet (marked by A in stage III) as a function of reaction time.

[1] W.T. Yuan, J. Yu, H.B. Li, Z. Zhang, C.H. Sun and Y. Wang, Nano Research 10(2), 397â&#x20AC;&#x201C; 404 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P12

Session: In Situ Nanoscale Microscopy of Processes Direct visualization of oxidation and reduction of FeO/Au(111) studied by time-resolved STM Yijia Li*1, Jeppe Vang Lauritsen1.

1

Interdisciplinary Nanoscience Center (iNANO), Aarhus University, DK-8000 Aarhus C, Denmark *E-mail: Yijia.Li@inano.au.dk Keywords: FeO, oxidation, reduction, STM. Iron oxide (FeO) is of significant interest due to its catalytical reactivity in several reactions such as CO oxidation and water-gas shift reaction [1], and as an intermediate product for the Fe-based Fischer-Tropsch catalysts [2]. The nature and the reaction mechanisms of the active sites are fundamental questions to heterogeneous catalysis. Therefore, scanning tunneling microscopy (STM) which is capable to identify the active sites at atomic scale level is employed. Here we studied the oxidation and reduction of the Au(111)-supported FeO islands using in-situ STM measurements recorded during oxygen or hydrogen exposure. It is found that the edges of FeO islands play an essential role for incorporating additional O atoms and O adatoms form triangular bright features assigned to the O adatom defects. The excess O atoms are in different coordination with lattice O atoms and can be removed by molecular hydrogen dosing. On the other hand, upon hydrogen exposure, the FeO islands are reduced with hydroxyls formed initially on the surface and vacancy defect loops created afterwards.

These O vacancy defects can be refilled by oxygen dosing. The reversible structural changes of the FeO islands depending on whether they are in oxidizing or reducing atmosphere are observed dynamically by the time-resolved STM movies and enable us to distinguish the active sites in redox reactions.

[1] Herzing, A.A., et al., 2008. 321(5894): p. 1331-5.

[2] de Smit, E. and B.M. Weckhuysen, Chem Soc Rev, 2008. 37(12): p. 2758-81.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P13

Session: In Situ Nanoscale Microscopy of Processes Investigating the reduction process of doped and undoped Ceria nanoparticles with in-situ TEM Annett Thøgersen*1, Kathrin Michel2,3, Matthias T. Elm2,3, T. Brezesinski4, and Truls Nordby5.

1

SINTEF Industry, Materials Physics, Forskningsveien 1, 0314 Oslo, Norway.

Institute of Physical Chemistry, Justus Liebig University Giessen, Heinrich-Buff-Ring 17, D-35392 Giessen, Germany 3 Center for Materials Research, Justus Liebig University Giessen, Heinrich-Buff-Ring 16, D-35392 Giessen, Germany 4 Institute of Nanotechnology, Karlsruhe Institute of Technology, Hermann-von-HelmholtzPlatz, D-76344 Eggenstein-Leopoldshafen, Germany 5 Centre for Materials Science and Nanotechnology, Department of Chemistry, University of Oslo, FERMiO, Gaustadalléen 21, NO-0349 Oslo, Norway *E-mail: annett.thogersen@sintef.no Keywords: Cerium, ETEM, EELS, Core-loss, Batteries.

2

Ceria (CeO2) nanoparticles have shown great potential for use in solid oxide fuel cells, oxygen sensors, oxygen separation membranes, and as catalyst material owing to their high electronic and oxygen ion conductivity and high chemical stability [1,2]. The particles exhibit high and reversible oxygen storage capacities, carried out by changing the valence state of Ce from 3+ to 4+, at intermediate temperatures (500 – 800 oC). Doping CeO2 with elements such as Pr and Tb, has shown to enhance its redox properties [1]. A change in valency during oxidation or reduction can be mapped on the nanoscale using the fine structure of the Ce-M4,5 edge with electron energy loss spectroscopy (EELS) [2].

In our work, we have mapped the change in valency during reduction and oxidation on nanopowder of pure CeO2 and mesoporous thin films of the solid solution ZrO2-CeO2 [3], in vacuum using a Protochips fusion heating holder and with a Protochips atmosphere holder, using an aberration-corrected FEI Titan G2 60-300 microscope with a Gatan GIF Quantum 965 EELS Spectrometer. Our results show that in high vacuum (10-9 Pa) the CeO2 nanoparticles have a layer of Ce3+ at the top surface, while the core remains as Ce4+. However, in the atmosphere holder at low vacuum conditions (0.5 atm), this surface layer was not observed.

Incorporation of Zr4+ in mesoporous CeO2 increased the reduction of the particles. At room temperature in high vacuum conditions, all of the ceria nanoparticles had been reduced to Ce3+.

Figure 1: Mapping the 3+ (red) and 4+ (blue) core loss EELS peak of Ce during in-situ heating in vacuum.

[1]: Anita M. D’Angelo and Alan L. Chaffee. ACS Omega, 2, 2544−2551 (2017) [2]: Stuart Turner, Sorin Lazar, Bert Freitag, Ricardo Egoavil, Johan Verbeecka Stijn Put, Yvan Strauvend and Gustaaf Van Tendeloo. Nanoscale, 3, 3385–3390 (2011) [3]: Hartmann, P.; Brezesinski, T.; Sann, J.; Lotnyk, A.; Eufinger, J. P.; Kienle, L.; Janek, J. ACS Nano 2013, 7, 2999−3013.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark B-P14

Session: In Situ Nanoscale Microscopy of Processes Visualizing nanoparticle self-assembly in solution using in situ TEM Utkur M. Mirsaidov 1Centre for Bioimaging Sciences and Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore 117543 2Department of Physics, National University of Singapore, 2 Science Drive 3, Singapore, 117551 3Centre for Advanced 2D Materials and Graphene Research Centre, National University of Singapore, 6 Science Drive 2, Singapore 117546. *E-mail: mirsaidov@nus.edu.sg Keywords: in situ TEM, nanoparticles, self-assembly Self-assembly of nanoparticles (NPs) is an important bottom-up nanofabrication method in which NPs spontaneously organize into ordered superstructures. The collective behavior of NPs in assemblies leads to optical, electronic, and mechanical properties that are distinct from those of the bulk materials and individual NPs. NP self-assembly within solutions is driven by intermolecular forces between NPs and external forces. However, owing to experimental challenges associated with imaging nanoscale dynamic processes in liquids, the interaction and assembly mechanisms of NPs are poorly understood.

Here, using direct in situ TEM imaging in liquid phase [1], I will describe the role of intermolecular forces, such as van der Waals, electrostatic forces, solvation, capillary, and other interactions in NP self-assembly in solution [2]. For example, I will show how the balance between the repulsive hydration force and attractive van der Waals (vdW) force for interacting NPs regulates their attachment dynamics [3]. I will also highlight how chemical functionalization of NPs can guide their self-assembly and the different pathways through which these NPs assemble [4-6].

Moreover, I will share our recent results on nanoscale elastocapillary effects that lead to collapse and aggregation of vertical nanowire arrays. I will describe the pathways through which drying, deflection, and adhesion of nanowires occurs and show that the formation of liquid nanodroplets is a common intermediary in the nanoscale pattern collapse.

Our findings the role of solvent-mediated physical and chemical forces in the assembly of nanoscale materials highlight the importance of direct nanoscale observation in uncovering previously unknown intermediate states that are pivotal for synthesis and selfassembly.

[1] H. Zheng, R. Smith, Y. Jun, C. Kisielowski, U. Dahmen, A. P. Alavisatos, Science 324 (2009), p.

1309.

[2] S. F. Tan, S. W. Chee, G. Lin, U. Mirsaidov, Accounts of Chemical Research 50 (2017), p.1303.

[3] U. Anand, J. Lu, N. D Loh, Z. Aabdin, U. Mirsaidov, Nano Lett. 16 (2016), p. 786.

[4] S. F. Tan, U. Anand, and U. Mirsaidov, ACS Nano 11 (2017), p.1633.

[5] G. Lin, S. W. Chee, S. Raj, P. Kral, and U. Mirsaidov, ACS Nano 10 (2016), p. 7443.

[6] S. F. Tan, S. Raj, G. Bisht, H. Annadata, C. A Nijhuis, P. Kral, U. Mirsaidov, Advanced Materials (2018) (DOI: 10.1002/adma.201707077).

Acknowledgments: This work is supported by Singapore National Research Foundation (NRF-CRP162015-05).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark D-I1

Session: In Situ Nanoscale Microscopy of Processes Measuring And Controlling Free-Standing Two-Dimensional Materials Ursula Ludacka*1, Mohammad R. A Monazam1, Toma Susi1, Christian Rentenberger1, Jannik C. Meyer1 and Jani Kotakoski1.

1

Faculty of Physics, University of Vienna, Austria. *E-mail: ursula.ludacka@univie.ac.at

Keywords: transmission electron microscopy, electron diffraction, two-dimensional materials, corrugations, mechanical strain.

In this work, we show through transmission electron microscope (TEM) and atomistic simulations that the non-flatness of free-standing graphene, hBN, and MoS2, as well as their heterostructures varies depending on the material. Out of the studied materials, graphene and hBN appear equally corrugated whereas MoS2 is rather flat. For the heterostructures, the overall shape is determined to a large extent by the stiffer of the two materials. In addition to measuring the out-of-plane shape, we can also control it in situ in one direction using a stretching holder. For these experiments, we glued the samples, transferred onto gold TEM grids with a perforated amorphous carbon film, onto the holder and applied mechanical strain with small incremental steps to avoid breaking the film during the experiment. Figure 1 shows the results of one such study. The deviation from circular symmetry of the diffraction pattern and the shape of the individual diffraction spots give us insight onto the strain in the material and its out-of-plane shape, respectively. Our results show that this simple method can be used to completely flatten the 2D materials in the direction of the applied force. At this point, the material exhibits an aligned set of one-dimensional corrugations. After the structure has been flattened, continuous mechanical deformation leads to a measurable strain in the structure.

Figure 1: Evolution of the diffraction pattern of graphene and individual diffraction spots during a straining experiment. Diffraction pattern recorded at different stages of the experiment (left) at the beginning, (middle) towards the end and (right) at the end. All shown patterns were recorded at sample tilt ι = 21°. The dashed lines show the approximate tilt axis and the overlaid hexagons highlight the first set of diffraction peaks. The panels on the right show a zoom-in of the indicated diffraction spots in false color.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark D-O1

Session: In Situ Nanoscale Microscopy of Processes In situ electron energy-loss spectroscopy for nanoscale optical devices Søren Raza*1 1

Department of Micro and Nanotechnology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark. *E-mail: sraz@nanotech.dtu.dk Keywords: nanophotonics, plasmonics, electron energy-loss spectroscopy, microelectromechanical systems.

Abstract: One of the most intriguing properties of surface plasmons is their ability to enhance the optical nearfield. This effect has given rise to numerous applications, including near-field optical microscopy, single molecule SERS, and bio-molecular sensors. Gapplasmon resonances in a metallic nanoparticle dimer is a prime example of such field confinement as it employs near-field coupling of two resonant antennas to further enhance the field intensity. Capacitive charge accumulation on the neighboring particle interfaces concentrates the electric field energy into the volume (gap) between the particles. As the particle spacing is reduced, the field intensity and the resonant frequency of the gap-plasmon mode become more sensitive to gap size variation. This high sensitivity to geometry renders the metallic nanoparticle dimer as a promising platform for actively tunable optical nanoantennas.

To achieve maximum field concentration and tunability of the resonance frequency, the gap size should be reduced to the ultimate limit. Here, we demonstrate dynamic electromechanical control over the coupling of a gold nanodisk dimer, and use this to systematically study the evolution of gap-plasmon resonances all the way to sub-nanometer-sized gaps. The device is prepared on a transmission electron microscopy (TEM) membrane, which allows us to apply the electrical actuation inside the microscope. By combining EELS with in situ electrical actuation, we can follow the evolution of the gap size and optical properties with unprecedented spatial and spectral resolution. We dub this new technique in situ EELS, which may become a critical tool for accessing the performance of future optical devices.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark D-O2

Session: In Situ Nanoscale Microscopy of Processes Organic ice resist lithography with an environmental TEM Anna Elsukova*1, Anpan Han1, and Marco Beleggia1.

1

DTU Danchip/Cen, Technical University of Denmark, 2800 Kongens Lyngby, Denmark. *E-mail: annaels@dtu.dk Keywords: Environmental TEM, nanofabrication, lithography, organic ice resists.

In this work we use an environmental transmission electron microscope (ETEM) to investigate the resolution limits of Organic Ice Resist Lithography (OIRL) [1]. OIRL is a novel one-step method for patterning nanostructures. Fig 1(a) outlines the general principle of OIRL. First, the organic vapor condenses into a thin layer of ice on the substrate, which is held at cryogenic temperature. Then, the ice layer is exposed to the scanning electron beam. After beam exposure, the substrate is heated to room temperature and unexposed ice sublimates. The areas exposed to the electron beam are non-volatile and remain on the substrate.

The size of the generated pattern depends on several factors, one being the illumination characteristics. To minimize the instrumental limitation on the patterning resolution imposed by a broad spot, we used the optics offered by an ETEM operated at 80 kV in scanning mode.

We used simple linear hydrocarbons (N-alkanes) with different molecular weights as precursors. After adjusting deposition and beam exposure parameters, we patterned sub-10 nm features (Fig 1(b)). The experiments revealed that the feature size depends on the precursor molecular weight. Coupling this result with the experimental contrast curves of each precursor (exposed thickness vs. dose) led to a model capable of reproducing the observed line-width vs. molecular weight trends, which is an essential step towards developing an understanding of the physics behind OIRL. We will also discuss a side-effect observed during our experiments, which could provide a new avenue to study beam-induced charging. Fig 1(c) shows three patterns obtained by scanning the electron beam with different dwell time per pixel. The discontinuities in the patterns are attributed to the sample jumps due to the accumulation and dissipation of charge.

Figure 1(a) General principles of OIRL. (b) Patterned lines on hydrocarbon organic ice. (c) Areas patterned for various beam dwell time per pixel [1] W. Tiddi, A. Elsukova, H.T. Le, P. Liu, M. Beleggia and A. Han, Nano Letters 17, 7886â&#x20AC;&#x201C; 7891 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark D-O3

Session: In Situ Nanoscale Microscopy of Processes: Fast Brownian Dynamics of Nanoparticles Observed in Liquid Phase Scanning Transmission Electron Microscopy Murat Nulati Yesibolati*,1, Hongyu Sun1, Kim I. Mortensen1, Sofie Tidemand-Lichtenberg1, Anders Brostrøm Bluhme1, Kristian Mølhave1 1

Department of Micro- and Nanotechnology, Technical University of Denmark, 2800 Kongens Lyngby, Denmark *E-mail: nuye@nanotech.dtu.dk

Keywords: Brownian motion, diffusion, liquid, scanning transmission electron microscopy Liquid phase transmission electron microscopy (LPTEM) allows investigating nanoparticle nucleation, growth and dynamics with high spatial and temporal resolution [1]. However, the nanoparticles near the solid-liquid interface in the LPTEM show different dynamics, specifically, 3-7 orders of magnitude smaller diffusion coefficients were typically reported compared to that of Brownian motion in bulk water [2]. The mechanisms behind the anomalously slow motion are not yet clear[3,4]. The nanoparticle dynamics can be dominated by the electron beam modified solid-liquid interface at the encapsulating silicon nitride membrane (Fig 1a), and given the unknown cause of the radically modified behavior, this could raise concerns of how much results on particles on the membranes can be generally trusted as representing ex-situ results.

Therefore, in this study, we studied Brownian motion of Au nanoparticles freely moving in the LPTEM in STEM mode as shown in Fig. 1a and b, and measured the translational diffusion coefficient using single particle tracking. For the first time, we report a diffusion coefficient matching that of free Brownian motion ( Fig 1c and d), which establishes LPTEM in STEM mode as a reliable tool with high spatiotemporal resolution for measuring dynamics of nanoparticles in their native environment.

a)

c)

d)

10

Time

MSD ( m2 )

Y ( m)

8 6 4

3

2

1.73 e-/(Å2×s) 0.89 e-/(Å2×s) 0.51 e-/(Å2×s) 0.25 e-/(Å2×s) 0,14 e-/(Å2×s) OM data

1

2 0 0

b)

4

2

4

6

X ( m)

8

10

0 0,0

0,1

0,2

0,3

0,4

0,5

0,6

Time (s)

0.5 µm

Figure 1: a) Schematic view of STEM imaging in a LPTEM; b) STEM image sequence of freely moving nanoparticles at electron flux 0.51 e-/(Å2×s), in 70 wt% Glycerol/water solution.

Frame interval time is 30 ms. For illustration purpose, the moving nanoparticles are marked with circle line; c) Trajectories of three randomly selected Au particles at the electron flux 0.51 e-/(Å2×s). Time interval is 30ms; d) Mean square displacement (MSD) vs. Δt.

[1] F.M. Ross et al, Liquid Cell Electron Microscopy, Cambridge: Cambridge University Press. (2016).

[2] L.R. Parent, E. Bakalis et al, Accounts of Chemical Research 51, 3-11 ( 2018).

[3] S.W. Chee, Z. Baraissov et al, Journal of Physical Chemistry C 120, 20462-20470 (2016).

[4] A. Verch, M. Pfaff , N. de Jonge, Langmuir 31, 6956-6964 (2015).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark D-O4

Advances in electron spectroscopy: techniques, instrumentation and applications Nanooptics in the electron microscope M. Kociak*1, L. H. G. Tizei1, H. Lourenço-Martins1, A. Campos1, D. Pabitra1 S. Meuret1, M. Tencé1, J. D. Blazit1, X. Li1, A. Gloter1, A. Zobelli1 and O. Stéphan1 1

Laboratoire de Physique des Solides, Université de Paris Sud, Orsay, France. *E-mail: Mathieu.kociak@u-psud.fr Keywords: EELS, cathodoluminescence, STEM, plasmons, excitons.

The use of electron spectroscopies, especially in the scanning transmission electron microscope (STEM), for studying nanooptics has exploded in the last 10 years. STEMCathodoluminescence (CL), STEM electron energy loss spectroscopy (EELS) and more recently electron energy gain spectroscopy (EEGS) are rapidly evolving, fueled by many instrumental and theoretical developments. Here, we will review some of these instrumental developments in the three fields that we participated in recently.

We will present recent advances in STEM-CL made in our lab, especially in the field of quantum optics 1,2. Switching to STEM-EELS, we will present results on plasmons in sub-2 nm silver nanoparticles, a domain of intense research at the moment 3 that were made possible thanks to an optimization of signal to noise ratio on a ~ 300 meV resolution NION USTEM. We will present the expected improvement when switching to a recently acquired sub-20 meV resolution monochromated NION STEM. Finally, first attempts to obtain EEGS, usually performed with a pulsed gun 4,5, but here without a pulsed gun, will be presented.

1.

2.

3.

4.

5.

Tizei, L. H. G. & Kociak, M. Spatially Resolved Quantum Nano-Optics of Single Photons Using an Electron Microscope. Phys. Rev. Lett. 110, 153604 (2013).

Bourrellier, R. et al. Bright UV Single Photon Emission at Point Defects in h-BN. Nano Lett. 16, 4317–4321 (2016).

Raza, S. et al. Blueshift of the surface plasmon resonance in silver nanoparticles studied with EELS. Nanophotonics 2, 1–8 (2013).

Barwick, B., Flannigan, D. J. & Zewail, A. H. Photon-induced near-field electron microscopy. Nature 462, 902–906 (2009).

Feist, A. et al. Quantum coherent optical phase modulation in an ultrafast transmission electron microscope. Nature 521, 200–203 (2015).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-I1

Session: Advances in electron spectroscopy: techniques, instrumentation and applications Cathodoluminescence measurements with an EDS detector Turkka Salminen*1,Paloma Del Cerro2, and Laeticia Petit2.

1

Tampere Microscopy Centre, Tampere University of Technology, Korkeakoulunkatu 3, 33720 Tampere, Finland.

2 Photonics Laboratory, Tampere University of Technology, Finland. *E-mail: turkka.salminen@tut.fi Keywords: Energy Dispersive X-ray Spectroscopy, Cathodoluminescence

Cathodoluminescence (CL) is a useful tool for the study of light emitting materials and defects in e.g. semiconductors, but CL detectors are far less common in SEMs than EDS detectors.

Typical EDS detectors are sensitive to light and their suitability for CL-measurements has been shown using the intensity of a light-induced low energy peak emerging in suitable conditions.[1] Here we present another approach using shifts of the X-ray peaks of the known elements in the luminescent material. The influx of light on the EDS detector creates extra charges leading to a leakage current-like signal. By adjusting the processing time and dwell time suitably the light induced charge is observed as peak shifts relative to the light intensity.

The combined CL and EDS analysis is demonstrated on glasses doped with persistent luminescent microparticles.

[1] P.F. Smet, J.E. Van Haecke and P. Poelman, Journal of Microscopy 231, 1â&#x20AC;&#x201C;8 (2008).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-O1

Session: Advances in electron spectroscopy: techniques, instrumentation and applications: Probing Three Dimensional Magnetic Information using Electron Vortex Beams with Nanometre-Scale Depth Resolution Devendra Singh Negi*1, Lewys Jones2, Juan-Carlos Idrobo3 and Jan Rusz1 1

Department of Physic and Astronomy, Uppsala University, Sweden.

Advanced Microscopy Laboratory, Centre for Research on Adaptive Nanostructures and Nanodevices (CRANN), Dublin 2, Ireland.

3 Center of Nanophase Materials Sciences, Oak Ridge National Laboratory, Oak Ridge, USA. *E-mail: dev.snegi1@gmail.com 2

Keywords: Electron Vortex Beam, Electron Magnetic Circular Dichroism, Magnetism.

Magnetic information at the nano-scale is essential for the growth of nano-technology. In this context, among the few available techniques i.e., SPEEM [1], XMCD [2]; EMCD is a promising approach [3]. However, all the techniques as yet limited to the lateral dimension.

Few efforts are paid to measure the depth sensitive imaging information in TEM [4]. We propose a three dimensional magnetic measurement method using EMCD in the transmission electron microscope. We study depth dependent EMCD of EVBs with varying defoci. A LaMnAsO crystal with 30 nm thickness is considered for simulation study. Inelastic cross sections of the Mn-L3 edge are calculated with the MATS.V2 software for probe convergence angles (α) of 30, 60 mrad respectively [5]. Figure 1, shows the depth dependent EMCD signal at different defocus values (- Δf nm) and a convergence angle of α = 60 mrad. The EMCD signal decreases with increasing defocus and thickness. Figure 2 shows the center of the magnetic signal intensity and the associated fullwidth at half-maximum (FWHM) as a function of defocus. The FWHM for 60 mrad is 1.8 nm. We thus propose that by collecting multiple spectrum images from same area by varying focus, it is possible to find the magnetic information in three dimensions.

Figure1: Crystal depth and defocus dependent EMCD signal.

Figure 2: Center of gaussian of EMCD intensity as a function of defocus.

[1] S. Heinze, et.al, Science 288, 1805–1808 (2000).

[2] W. Chao, et.al, Nature 435, 1210 (2005).

[3] P. Schattschneider, et.al, Nature 441, 486-448 (2006).

[4] J. G. Lozano, et.al, Phys. Rev. Lett. 113, 135503 (2014).

[5] J. Rusz, et.al, Ultramicroscopy 177, 20-25 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-O2

Session: Advances in electron spectroscopy: techniques, instrumentation and applications A systematic comparison of on-axis vs. off-axis transmission Kikuchi diffraction F. Niessen1,a, A. Burrows2,b, A. Bastos da Silva Fanta2,c 1

Danish Hydrocarbon Research and Technology Centre (DHRTC), Technical University of Denmark (DTU), Elektrovej, 2800 Kgs. Lyngby, Denmark

2

Center for Electron Nanoscopy (CEN), Technical University of Denmark (DTU), Fysikvej, 2800 Kgs. Lyngby, Denmark a

frannie@dtu.dk; bandrew.burrows@cen.dtu.dk; casf@cen.dtu.dk;

Transmission Kikuchi diffraction (TKD) is becoming an increasingly popular nanocharacterization technique, which conventionally operates on standard electron backscatter diffraction (EBSD) hardware and an electron-transparent sample. Fundenberger et al. recently introduced a detector setup that is optimized for TKD, consisting of a phosphor screen positioned below the sample normal to the incident beam, analogous to a bright field detector in transmission electron microscopy (TEM) [1]. As the acquisition occurs on the axis of the incident beam, TKD in this configuration is termed “on-axis” TKD, compared to conventional “off-axis” TKD [2].

The on-axis transmission Kikuchi diffraction (TKD) technique was systematically compared with conventional off-axis TKD [3]. The effect of experimental parameters on the appearance of on-axis and off-axis Kikuchi patterns was investigated. It was found that, in contrast to offaxis TKD, on-axis TKD is more sensitive to changes in beam current and beam energy and less sensitive to changes in working distance and detector distance. The most significant advantage of on-axis TKD over off-axis is enhanced pattern intensity, which allows reduction of the beam current or an increase in the acquisition rate. The physical and effective lateral spatial resolution were measured with detector-typical parameters and were found to differ, where off-axis TKD is more sensitive to beam drift. The Hough transform was found to operate more robustly on on-axis TKD patterns. When using an on-axis detector, the Kikuchi pattern includes the transmitted beam, which leads to a bright spot in the center of the pattern.

It was found that the bright spot caused by the transmitted beam in on-axis TKD did not noticeably disturb the Hough transform. In the case where the major zone axis coincided with the bright spot of the transmitted beam, additional zone axes in the periphery of the Kikuchi pattern could be detected.

[1]

J.J. Fundenberger, E. Bouzy, D. Goran, J. Guyon, H. Yuan, A. Morawiec, Orientation mapping by transmission-SEM with an on-axis detector, Ultramicroscopy. 161 (2016) 17–22. doi:10.1016/j.ultramic.2015.11.002.

[2] H. Yuan, E. Brodu, C. Chen, E. Bouzy, J.-J. Fundenberger, L.S. Toth, On-axis versus off-axis Transmission Kikuchi Diffraction technique: application to the characterisation of severe plastic deformation-induced ultrafine-grained microstructures, J. Microsc. 0 (2017) 1–11. doi:10.1111/jmi.12548.

[3] F. Niessen, A. Burrows, A. Bastos da Silva Fanta, A systematic comparison of on-axis and off-axis transmission Kikuchi diffraction, Ultramicroscopy. (2017). doi:10.1016/j.ultramic.2017.12.017.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-O3

Session: Advances in electron microscopy: techniques, instrumentation and applications Organic Ice Resists William Tiddi1, Anna Elsukova1, Ding Zhao1, Marco Beleggia1, Anpan Han1*.

1

Danchip/CEN, Technical University of Denmark, Kgs. Lyngby Denmark E-mail: anph@dtu.dk

Keywords: electron beam lithography, organic ice, instrumentation, cryostage.

Electron-beam lithography (EBL) is the backbone technology for patterning nanostructures and manufacturing nanodevices. It involves processing and handling synthetic resins in several steps, each requiring optimization and dedicated instrumentation in cleanroom environments. Here, we show that simple organic molecules, e.g. alcohols, condensed to form thin-films at low temperature demonstrate resist-like capabilities for EBL applications and beyond. The entire lithographic process takes place in a single instrument, and avoids exposing chemicals to the user and the need of cleanrooms. Unlike EBL that requires large samples with optically flat surfaces, we patterned on fragile membranes only 5nm-thin, and 2 x 2 mm2 diamond samples. We created patterns on the nm to sub-mm scale, as well as three-dimensional structures by stacking layers of frozen organic molecules. Finally, using plasma etching, the organic ice resist (OIR) patterns are used to structure the underlying material, and thus enable nanodevice fabrication.

This is the first time we present out results at an electron microscopy conference. We will also show newest results beyond our paper [1].

Figure: OIR patterning compared to other e-beam based techniques. (a) EBL, (b) FEBID. (c) IL, (d) OIR â&#x20AC;&#x201C; A vapour of a simple organic compound is first condensed onto the cooled sample to form a uniform layer. Its interaction with the e-beam locally modifies its chemical composition resulting in a non-volatile product. When the sample is heated to room temperature the unexposed OIR sublimates while the exposed patterns are stable, enabling ambient downstream processing.

[1] W. Tiddi, A. Elsukova, H. T. Le, P. Liu, M. Beleggia, A. Han, Nano Letters, 2017, 17, 7886

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-O4

Session: Advances in electron microscopy: techniques, instrumentation and applications Ice resists for 3D electron-beam processing: instruments in Denmark and China Ding Zhao*1, William Tiddi1, Anna Elsukova1, Marco Beleggia1, Min Qiu2,3, Anpan Han1.

1

2

Danchip/CEN, Technical University of Denmark, Kgs. Lyngby, Denmark College of Optical Science and Engineering, Zhejiang University, Hangzhou, P.R. China 3 Westlake University, Hangzhou, P.R. China. *E-mail: dizhao@dtu.dk Keywords: electron beam lithography, organic ice, instrumentation, cryostage.

Water and organic vapors condensed into thin layers of ice on the surface of a cold substrate can be exposed with an electron beam to create resist patterns for lithography applications.

The entire spin- and development-free lithography process usually requires a single custom instrument.

We report the design, material choice, implementation and operation of two instruments; one at Zhejiang University (ZJU), China, and one at DTU, Denmark. Both are based on scanning electron microscopes fitted with an electron beam control system that is normally used for electron beam lithography. The microscopes are equipped with gas injection systems, liquid nitrogen cooled cryostages, temperature control systems, and load-lock chambers. The Zhejiang Universty instrument can also deposit metals at cryogenic temperature for pattern transfer. By stacking nanoscale patterns made in organic ice, 3D nanostructures are created through complementary cyclic condensation, exposure and sublimation processes.

Figure: Organic ice resist processing sequence and AFM images of fabricated 3D structures (left). DTU (center) and ZJU instrument (right).

[1] W. Tiddi, A. Elsukova, M. Beleggia, and A. Han, Microelectronic Engineering 192, 38-43 (2018).

[2] W. Tiddi, A. Elsukova, H. T. Le, P. Liu, M. Beleggia, and A. Han, Nano Letters 17, 78867891 (2017).

[3] A. Han, A. Kuan, J. Golovchenko, and D. Branton, Nano Letters 12, 1018-1021 (2012).

[4] A. Han, J. Chervinsky, D. Branton, and J. A. Golovchenko, Review of Scientific Instruments 82, 065110 (2011).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-P1

Session: Advances in Electron Spectroscopy: Techniques, Instruments and Applications Superseding CCD with CMOS Technology in EBSD – Massively Increased Speed, Sensitivity and Resolution in a Single Detector Keith Dicks*1, Angus Bewick1.

1

Oxford Instruments NanoAnalysis, Halifax Rd, High Wycombe, Bucks HP12 3SE. *E-mail: keith.dicks@oxinst.com Keywords: Binning, solution rates, sensitivity, speed.

The CCD (Charge Coupled Device) commonly used as the image sensor in EBSD detectors is a serial device – the pixels are read out sequentially. Pixel ’binning’ is used to increase frame rates, and to boost the signal level in each read pixel. A high-resolution, high-sensitivity detector may have a full resolution of 1344x1024 pixels; operated at 8x8 binning, it may achieve a max. solution rate of ~100pps at >100pA. Conversely, High-speed EBSD detectors use VGA format CCD sensors (640x480 pixels, at max. ~200Hz). Again, binning increases frame rates, whilst also improving the low electron dose detection limit (because the read-out noise decreases relative to the signal in the binned 'super pixel'). To achieve the highest solve rates (~1600pps at >12nA), 16x16 binning is required which has two very significant disadvantages: reduction of the EBSP resolution to 40x30 pixels; and reduction of the maximum signal to noise ratio (SNR) before image saturation (owing to the limited charge capacity of the CCD's output node). At such extreme binning levels, accurate band detection and indexing is compromised, and angular precision of orientation measurements is reduced.

With the ‘massively parallel’ device architecture of a CMOS (Complementary Metal Oxide Semiconductor) image sensor, many outputs are read simultaneously, greatly increasing frame rates without the need for pixel binning. Thus CMOS can offer significant advantages over CCD for EBSD detectors – higher speed at higher resolution, and with higher SNR. Compared to the best high-speed CCD detector, a CMOS can achieve twice the speed for the same electron dose, with four times the resolution: >3000pps at >12nA and 156x128 pixels.

CMOS offers high speed with high indexing rates and high angular precision, even with lowcontrast EBSPs where the low SNR of CCD fails. CMOS also out-performs high-resolution, high-sensitivity CCDs: 250pps at >250pA and 1244x1024 pixels. See Figure 1. With CMOS technology, a single EBSD detector can now encompass all applications, exceeding the performance of both high-resolution and high-speed CCD detectors, coupled with significantly higher sensitivity.

Figure 1: 1244 x 1024, 250Hz at 250pA vs. binned to 156 x 128 pixels, 3000Hz, 12nA

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-P2

Session: Advances in electron spectroscopy: techniques, instrumentation and applications Cryo EM workflows for Single Particle Analysis and Tomography of hydrated, intact cells Max Maletta*1, Wim Voorhout, M. Storms, Gijs van Duinen, J. Lengyel, M. Vos and B. Lich.

1

ThermoFisher Scientific, Achtseweg Noord 5, 5651 GG Eindhoven, The Netherlands *E-mail: max.maletta@thermofisher.com Keywords: Cryo-EM, Integrative Biology, Tomography, In situ structural biology.

A new frontier exists in unraveling interactive biological and biochemical processes and pathways at the macromolecular level. Of critical importance is the three-dimensional visualization of macromolecular structures and molecular machines in their native functional state. Three techniques play a major role, NMR, XRD and Cryo-TEM. Nuclear magnetic resonance (NMR) has the capability to study specific protein domains or fragments and their functional role in protein folding and dynamics and in ligand binding whereas X-Ray crystallography (XRD) allows visualizing high-resolution but more static 3D structures of apo and liganded proteins, mainly in a monomeric or dimeric state after crystallization. To unravel more physiologically relevant situations however, it is essential to visualize multimeric complexes in their tertiary and quaternary state and their interaction with other complexes. Cryo-TEM applications like single particle analysis one can visualize multimeric complexes. In this so-called translational methodology, cryo-TEM thus provides complementary information to NMR and XRD that can be crucial for a detailed structural analysis for a better understanding of the mechanism of the physiologically relevant complex.

Latest developments in the cryo-TEM workflow have brought the 3 major structural biology technologies closer together. Now, finally, a continuum has been reached on all important aspects with regards to resolution and macromolecular scales which allows for the full deployment of the combination of these technologies.

We will discuss the future of structural biology based on the latest developments of the FEI workflow and its components.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-P3

Session: Advances in electron microscopy: techniques, instrumentation and applications Novel Automation Scripting for SEM & DualBeam D. Phifer*1 1

Thermo Fisher Scientific, Eindhoven, The Netherlands. *E-mail: daniel.phifer@thrermofisher.com

Keywords: scripting, automation, SEM, DualBeam, in situ.

Repetitive electron microscopy tasks that could be automated are often not due to the microscope/instrument manufacturer not allowing control of these systems. Manufacturers chose the most commercially viable routines to sell as dedicated software such as "Auto Slice and View" or "Auto TEM" and these applications are quite successful for high volume applications. Unfortunately, many applications that are routine and repetitive are extremely specific to real research needs and there is a real desire for control and automation solution for SEM and DualBeamâ&#x201E;˘ systems to meet this need. Python appears to be the premier open source scripting language for scientific research so Thermo Fisher Scientific has developed a Python interface library, "AutoScript", to interfaces the SEM and DualBeam platforms with modern operating systems to enable simple automation routines to be constructed which can do a variety of routine and non-routine unique tasks.

The AutoScript API allows automation routines to be constructed which can do a variety of non-routine tasks as well and decision trees can be integrated for unattended operation such as in Fig.1. Combining control of most all microscope functions including setting and reading values, calling auto functions, saving images and storing values with other existing open source Python scripts can be supplemented with image matching, data graphing, automated response and feedback loops to create novel performance. Many nice examples of custom scripts have already been created that adjust settings with a feedback loop, extract and graph in situ data, and perform image capture automatically at specified positions. The availability of so many Python based routines in the open source environment allows various components to be integrated/combined for specific purposes. With the ability to control both the SEM and FIB columns, GIS, Manipulator, and other accessories, the AutoScript interface allows writing advanced automation for most tasks. Python libraries exist for image matching, data display and data export that complete the suite of capabilities needed for custom automation of systems on Windows 7 and higher.

Figure 1: Example of complex FIB patterning and subsequent SEM measurements

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark E-P4

Session: Novel applications for X-rays and SPM Using X-ray Imaging to Visualise the 3D Architecture of White Matter Mariam Andersson1,2, Hans Martin Kjer1,2, Vedrana Andersen Dahl2, Martin Bech3, Alexandra Pacureanu4, Anders Bjorholm Dahl2, Tim B. Dyrby1,2.

1

Danish Research Centre for Magnetic Resonance, Copenhagen University Hospital Hvidovre, Hvidovre, Denmark 2 Department of Applied Mathematics and Computer Science, Technical University of Denmark, Kongens Lyngby, Denmark 3 Department of Medical Radiation Physics, Clinical Sciences, Lund University, Lund, Sweden 4 European Synchrotron Radiation Facility, Grenoble, France *E-mail: maande@dtu.dk Keywords: x-ray phase contrast tomography, MRI, axon, white matter An understanding of the 3D microstructural environment in the brain is required to design improved biomarkers for clinical magnetic resonance imaging (MRI) diagnosis. This study aims to determine the 3D microstructural architecture of white matter and visualise relative trajectories of anatomical structures that are imaged by clinical MRI. We obtained the first ultra-high resolution 3D images of white matter samples at a sub-micrometer scale with X-ray phase contrast tomography at the European Synchrotron Radiation Facility (ESRF) in France.

Neurodegenerative diseases, such as Multiple Sclerosis, cause anatomical changes to axons and their surrounding myelin sheaths. Such changes can be measured with non-invasive clinical MRI. Since the MRI resolution is around 1-2 mm, and axon diameters range between 0.5-5 ď ­m, a biophysical model [1, 2] which assumes that axons are straight, aligned tubes is employed at the Danish Research Centre for Magnetic Resonance (DRCMR) to estimate axon diameters/densities. Hereby, clinically relevant information is averaged across many axons, and 3D structures like bending axons are not accounted for. High-resolution X-ray phase contrast tomography enables detailed imaging of these 3D structures and their environment.

X-ray phase contrast tomography (voxel size 75-100 nm) was performed at ID16A, ESRF, on osmium-stained white matter biopsies from a post-mortem monkey brain. Figure 1a. depicts a slice from a crossing fibre region; axons and vessels appear bright and tube-like, with the dark myelin border being axon-specific. Figure 1b shows a 3D visualisation of a selection of large axons and blood vessels.

The ultra-high resolution 3D images of white matter from X-ray phase contrast tomography contain detailed anatomical information that can be used to improve biophysical MRI models and increase their sensitivity to disease-induced microstructural changes.

a

b

Figure 1: a) 2D X-ray image depicting a white matter crossing fibre region. An axon, surrounded by a dark myelin sheath, is shown in yellow and a vessel is shown in red. Scale bar: 20 Âľm b) Crossing fibre region 3D view: large axons (yellow) and vessels (red).

[1] [2]

S. N. Jespersen, et al. Frontiers in Physics. doi:10.3389/fphy.2014.00028, (2014).

H. Lundell, et al. Magn. Reson. Med., doi: 10.1002/mrm.25211, (2015).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-I1

Session: Novel applications for X-rays and SPM In situ Characterization of Catalysts: Combining X-ray and electron microscopy Christian Danvad Damsgaard *1,2, Sina Baier2,3, and Jan-Dierk Grunwaldt3,4.

DTU Danchip/Cen, Technical University of Denmark, Kgs. Lyngby, Denmark.

2 DTU Physics, Technical University of Denmark, Kgs. Lyngby, Denmark.

3 Institute for Chemical Technology and Polymer Chemistry, Karlsruhe Institute of Technology, Karlsruhe, Germany.

1

4

Institute of Catalysis Research and Technology, Karlsruhe Institute of Technology, Karlsruhe, Germany. *E-mail: christian.damsgaard@cen.dtu.dk

Keywords: In situ, Environmental TEM, Ptychography, Catalysis, X-ray microscopy Fundamental insight into structure-functionality relationships is required to develop and improve properties of heterogeneous catalysts. As catalysts may change their structure with respect to the environment, it is essential to investigate the catalysts under reaction conditions.

Furthermore, structural and compositional information have to be acquired on different length scales[1] and such in situ studies require dedicated complementary techniques. Traditionally, nanoscale imaging and spectroscopy of catalysts in a gaseous environment is performed in an environmental transmission electron microscope (ETEM). TEM gives insight in the atomic changes during reaction, however it is restricted to relatively low pressure (<~1 kPa) and a thin sample (<~100 nm)[2]. Spatially resolved information on the meso scale (50 nm–1 µm) can be obtained by X-ray microscopy, which enables in situ studies at both ambient and elevated pressure[3]. This contribution elucidates catalyst properties by combining X-ray and electron based microscopy.

One example highlighted combines X-ray imaging with ETEM studies of a bifunctional Cu/ZnO@zeolite core-shell catalyst for direct production of methanol[4]. The study reveals (Fig. 1) a stable core-shell interface at 250°C, although reduction of the Cu containing core material led to a shrinkage of the particles on the nanometer scale. At further heating to 350°C changes on the μm scale were observed.

The results underline the need for complementary techniques and highlight the potential of these for application in catalysis.

Figure 1: In situ ptychograms (phase contrast) of a thin slice of a Cu/ZnO@zeolite coreshell catalyst at room temperature in He, 250°C in H2, 250°C in O2, 350°C in H2, and 350°C in O2, respectively [4].

[1] [2] [3] [4]

J. D. Grunwaldt et al., ChemCatChem, vol. 5, no. 1, pp. 62–80, 2013.

T. W. Hansen et al., Science, vol. 294, no. 5546, pp. 1508–1510, 2001.

S. Baier et al., Microsc. Microanal., vol. 22, no. 1, pp. 178–188, 2016.

S. Baier et al., Microsc. Microanal., vol. 23, no. 3, pp. 501–512, 2017.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-O1

Session: Novel applications for X-rays and SPM Soft X-ray Imaging of Endothelial Cells and Their Glycocalyx Casper Hempel*1, Sergey Kapishnikov2, Thomas Lars Andresen1, and Klaus Qvortrup3.

1

Department of Micro- and Nanotechnology, Technical University of Denmark, Lyngby, Denmark 2 Niels Bohr Institute, University of Copenhagen, Copenhagen, Denmark.

3 Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark. *E-mail: casperhempel@gmail.com Keywords: endothelial cells, glycocalyx, soft x-ray imaging.

Endothelial cells are lining all blood vessels and form the interface between tissue and blood.

On the luminal side one finds a carbohydrate-rich matrix, which is termed the endothelial glycocalyx. It plays a key role in maintaining vascular function and is rapidly lost during various types of stimuli and pathology. Due to its biological importance and the lack of ideal visualisation techniques we aimed to improve the preservation and subsequent the imaging quality of the luminal endothelial glycocalyx.

Soft x-ray imaging allows for submicron resolution of hydrated, native state cells in the water window. In this study we performed cryo, soft X-ray imaging of two types of specimens: Human brain endothelial cells grown on gold grids and capillaries isolated from mouse brains.

Samples were preserved by high pressure freezing. By using this procedure plasma membranes, nuclei and intracellular organelles are visible. The glycocalyx did not possess enough contrast on its own to be resolved so cationized ferritin and lanthanides were added to resolve the glycocalyx. These agents are commonly used in transmission electron microscopy imaging of this structure. The obtained results are compared with data obtained using transmission electron microscopy of cryogenically preserved specimens of comparable type.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-O2

Session: Scanning Probe Microscopy Combining Ultra High Vacuum Scanning Tunneling Microscopy and Electrochemistry for Surface Studies of Model Catalysts Thomas Maagaard*1, Sebastian Horch1, and Ib Chorkendorff1.

1

SurfCat, Department of Physics, The Technical University of Denmark, Fysikvej 307, 2800 Kgs. Lyngby, Denmark. *E-mail: thomaa@fysik.dtu.dk Keywords: STM, UHV, electrochemistry, active sites, corrosion.

Understanding surface processes is important in understanding electrocatalytic reactions.

In order to characterise a given catalyst as close as possible to its state before and after testing it electrochemically we have combined an ultra high vacuum (UHV) chamber with an electrochemical setup. This makes it possible to avoid any potentially contaminating sample transfer steps.

The UHV chamber is equipped with a scanning tunneling microscopy (STM) which allows for atomic resolution of the surface of our catalyst making it possible to investigate effects related to catalytically active sites and corrosion phenomena.

We present two cases, firstly Pt(111) as it is the benchmark system in electrocatalysis, and secondly Cu(111) as copper has been shown great interest as an electrocatalyst within the CO and CO2 reduction communities. These will serve as a basis for discussing the capabilities and limitations of the method at present.

From left to right: 700 nm 700 nm STM image of a freshly prepared Pt(111); the first cyclic voltammogram to 0.9 V vs. RHE after immersion; representative corrosion cycles to 1.2 V vs. RHE; 700 nm 700 nm STM image of a corroded Pt(111) surface.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-O3

Novel applications for X-rays and SPM Detailed Surface Examinations of III-V Nanowires by Scanning Probe and X-ray Photoelectron Spectroscopy Techniques Sarah R. McKibbin*1, J. V. Knutsson1, S. Yngman1, J. Colvin1, A. Trojan1, Y. Liu1, Y.-P. Liu1, J. L. Webb1, K. A. Dick1, R. Timm1 and A. Mikkelsen1

1. Department of Physics and NanoLund, Lund University, Lund, Sweden *E-mail: sarah.mckibbin@sljus.lu.se Keywords: scanning probe microscopy, scanning tunneling spectroscopy, semiconductor nanowires, x-ray photoelectron spectroscopy.

III-V nanowires (NWs) are highly promising candidates for efficient optoelectronic, integrated circuit and photovoltaic devices.[1, 2] With typical diameters of 20 to 100 nm and lengths of several Îźm their structures they can contain complex heterostructures with a broad range of materials, doping, or crystal structures to choose from. As surface effects dominate over bulk properties in narrow devices, it is crucial to understand the electronic, structural and chemical makeup of such NWs along their length down with high precision to properly correlate these parameters to performance and allow for further optimization.

An enormous level of detail in NW can be revealed through multiple surface analysis methods such as scanning probe and photoelectron microscopy techniques,[3] which we can then combine with simultaneous electrical operation.[4] We present atomic force microscopy (AFM), scanning tunnelling microscopy (STM) and x-ray photoelectron spectroscopy (XPS) results GaAs NWs and correlate changing doping levels to structural changes along the wires and the formation of native oxides. We show preliminary results on individual GaAs and InAs NWs in a complete device architecture to investigations into atomic scale features on the NW surface with STM and scanning tunnelling spectroscopy (STS) with simultaneous electrical measurements.

We will then present results from InP NWs containing a p-n junction to detail the potential landscape along the NW surface and at the doping interface using kelvin probe force microscopy (KPFM) and STM/STS (Fig 1 a, b, c). In-operando measurements were also performed by taking chemical maps of an InP p-n junction NW using nanofocused XPS on a lithographically defined device to image the entire sample environment and probe chemical shifts on the surface (Fig 1 d) whilst the device is under forward and reverse bias.

[1] J. Wallentin et al., Science 339, 1057 (2013); [2] E. Lind et al., IEEE J. El. Dev. Soc. 3, 96 (2015).

[3] M. Hjort et al., ACS Nano 6, 9679 (2012); M. Hjort et al., ACS Nano 8, 12346 (2014); [4] J. L. Webb et al., Scientific Reports 7, 12790 (2017).

Figure 1: InP nanowire (a) STM overview (b) KPFM (c) STS across p-n junction (d) Device setup and nano-xps results of in-operando measurements.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-O4

Session: (Novel applications for X-rays and SPM) STM and XPS Studies of Titania-Ceria Mixed Oxide Thin Films Tao Xu*1, Stefan Wendt1, Jeppe V. Lauritsen1.

1

Aarhus University, Interdisciplinary Nanoscience Center, Gustav Wieds Vej 14, 8000, Aarhus C, (DENMARK). *E-mail: tao.xu@inano.au.dk Keywords: Ceria, Titania, oxide thin film, STM, XPS.

The reducible oxides, Titania (TiO2) and Ceria (CeO2) are among the most widely applied oxide materials in various catalytic applications, where they are used as support to disperse catalytically active species like metal particles or oxide species or as catalysts in their own right. It has been found that titania and ceria actively participate in the catalytic reaction cycle and can have significant influence on the overall performance of the catalyst. Although they have been separately studied extensively, comparably little is known about the mixed compound CexTi1-XO2. Recently, experimental and theoretical work suggested that mixed CeO-Ti phase is catalytically active for many reactions.[1] For instance, selective catalytic reduction (SCR) of NOX,[2] selective dehydration of methanol,[3] etc. It is proposed that Ce3+ is stabilized in this mixed oxide phase and the reducibility of such mixed oxide is improved.[4] However, there is still lack of atomic understanding on the Ce-O-Ti phase and its exceptional catalytic activity.

In this work, we follow a rigorous surface science approach to grow atomically welldefined CeO2(111) thin film on a Ru(0001) single crystal under ultrahigh vacuum (UHV) conditions. Scanning Tunnelling Microscopy (STM) and X-ray photo-electron spectroscopy (XPS) are applied to examine the topography and electronic properties of the CeO2(111) thin film. Secondly, we explore different strategies to mix Ti with CeO2. By sequential deposition or co-deposition and post annealing in O2 or UHV, we systematically examine the obtained mixed oxide phases by STM and XPS. Based on the structural understanding gained in the above two steps, we further explore the adsorption behaviour of NH3, NO and O2 on these mixed Ce-O-Ti phases by STM and XPS, in combination with temperature programmed desorption (TPD). The current research provides systematic and atomic understanding of the titania-ceria mixed oxide and contribute to the rational synthesis of novel titania-ceria mixed oxide for catalysis.

[1]A. R. Albuquerque, A. Bruix, I. M. G. dos Santos, J. R. Sambrano, F. Illas, The Journal of Physical Chemistry C, 118, 9677-9689(2014); A. R. Albuquerque, A. Bruix, J. R. Sambrano, F. Illas, The Journal of Physical Chemistry C, 119, 4805-4816(2015).

[2]T. H. Vuong, J. Radnik, J. Rabeah, U. Bentrup, M. Schneider, H. Atia, U. Armbruster, W. GrĂźnert, A. BrĂźckner, ACS Catalysis, 7, 1693-1705(2017).

[3]S. Agnoli, A. E. Reeder, S. D. Senanayake, J. Hrbek, J. A. Rodriguez, Nanoscale, 6, 800810(2014).

[4]A. C. Johnston-Peck, S. D. Senanayake, J. J. Plata, S. Kundu, W. Xu, L. Barrio, J. Graciani, J. F. Sanz, R. M. Navarro, J. L. G. Fierro, E. A. Stach, J. A. Rodriguez, The Journal of Physical Chemistry C, 117, 14463-14471(2013).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-P1

Session: Novel applications for X-rays and SPM A Hard X-ray Nanoprobe for Multi-Modal Analysis at Diamond Light Source Julia E. Parker*1, Fernando Cacho-Nerin1, and Paul D. Quinn1.

1

Diamond Light Source, Diamond House, Harwell Science and Innovation Campus, Didcot, Oxfordshire, U.K. *E-mail: julia.parker@diamond.ac.uk Keywords: x-ray Nanoprobe, correlative imaging, XRF, XANES, nano-XRD,

The hard X-ray nanoprobe beamline at Diamond is a new facility for nanoscale microscopy.The beamline operates over a 4.5â&#x20AC;? to 23â&#x20AC;?keV energy range with an emphasis on multi-modal analysis providing elemental mapping, speciation mapping by XANES, structural phase mapping by nano-XRD and imaging through differential phase contrast and ptychography. A flexible scanning system allows for fast acquisition and arbitrary scan paths, simple data acquisition software and the ability to process data in near real-time. I14 welcomed its first users in March 2017 for commissioning experiments with its first user run between Oct 2017 and Mar 2018. The beamline is in an optimization phase with new techniques and users tools rolling out over a two year ramp up period.

The I14 beamline facility co-located with a new national electron microscopy facility providing electron microscopy suites covering the physical [1] and life sciences [2]. This facility combines staff and expertise from a number of different areas allowing allow us to make exciting progress in sample preparation techniques and correlative x-ray and electron microscopy studies., including development of an in situ heating and gas flow sample environment to allow correlative TEM and x-ray nanoprobe studies.

The beamline complements electron and optical microscopy and enables new science in a number of areas spanning materials science, biology, engineering and earth science. Here we present the current status and future development plans of Beamline I14, and highlight results from successful experiments to date.

Figure 1: Schematic of beamline I14 [1] http://www.diamond.ac.uk/Science/Integrated-facilities/ePSIC [2] D.K. Clare et al., Acta Cryst. D 73, 488 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-P2

Session: Novel applications for X-rays and SPM High Frequency Cantilever Evaluations for High Speed Atomic Force Microscopy in Liquid Harpreet Singh BRAR, Müjdat BALANTEKIN. Department of Electrical and Electronics Engineering Izmir Institute of Technology, Izmir, Turkey. *E-mail: er.harpreetbrar@gmail.com

In life sciences, High-Speed Atomic Force Microscopy is now widely accepted as a dynamic event visualizer for numerous biological samples such as live cells, membrane lipids, ATPproteins, Enzymatic reactions, DNA-protein interactions, [1,2] etc. HS-AFM’s unique ability to observe surface topography of the samples with height data and with a resolution of up-to a single atom makes it a prominent tool in nanoscale measurements.

HS-AFM Imaging technique’s speed and response is limited by various factors including cantilever probes, operating environment, scanning techniques, etc. Cantilevers are indispensable and integral part of HS-AFM, thereby necessitating their own critical evaluations. As the cantilevers play essential role in HS-AFM Imaging, the analysis of various parameters such as resonance frequency, stiffness, and Q-factor of cantilevers is an active area of research. Distinct number of authors have reported difficulties in calibrating cantilevers in liquid environments. Thus, the evaluation of cantilever parameters in liquid environment is essential.

Figure 1: Finite element simulation of silicon cantilever showing third flexural mode (liquid environment). Inset: Full geometrical model.

The research work based on finite element simulations focuses on the effects of cantilever material and geometry on the higher eigenmode resonance frequencies and Q-factors of cantilevers used in high-speed atomic force microscopy.

[1] Kodera, Noriyuki, D. Yamamoto, R Ishikawa, and Ando. Nature 468, no.7320 (2010): 7276.

[2] Uchihashi, Takayuki, R. Iino, T. Ando, and H. Noji. Science 333, no. 6043 (2011): 755758.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-P3

Session: Novel applications for X-rays and SPM Real time imaging of microstructural transformations in bulk ferroelectrics Jeppe Ormstrup1, Magnus Christensen2, Ragnvald Mathisen2, Phil Cook3, Can Yildirim3, Henning Friis Poulsen1, Hugh Simons1 1 2

Department of Physics, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark Department of Physics, Norway University of Science and Technology, NTNU NO-7491 Trondheim, Norway 3 ESRF – The European Synchrotron, Avenue des Martyrs, 38000 Grenoble, France *E-mail: jepor@dtu.dk

Keywords: diffraction, microstructures, kinetics, dark field, reciprocal space map, strain maps Ferroelectrics are a broad class of functional materials with the ability to store electric charge, convert between electrical and mechanical work, and store digital memory states. Their functionality derives from the formation and dynamics of structural domains (i.e. twins), which nucleate, reorganize and annihilate under applied electric, thermal or mechanical loads.

Characterizing these processes remains a persistent challenge due to the wide range of length (nm to mm) and time (µm to minutes) scales over which they occur. Here we present the use of Dark-Field X-ray Microscopy for directly imaging and tracking individual domains in real time during phase transformations and external perturbations. We describe the methodology and its application to study electric-field induced phase transitions in barium titanate. This capability to quantitatively correlate the structural dynamics to external boundary conditions is a key requirement for formulating and validating multi-scale models that account for the full heterogeneity of ferroelectric materials.

[1] A.K. Tagantsev, L.E. Cross, J. Fousek, Domains in ferroic crystals and thin films, New York: Springer (2010) [2] H. Simons, A. King, W. Ludwig, C. Detlefs, W. Pantleon, S. Schmidt, F. Stöhr, I. Snigireva, A. Snigirev, H. F. Poulsen. Nat. Commun. 6, 6098 (2015).

[3] M.F. Horstmeyer, Integrated Computational Materials Engineering (ICME) for Metals: Using Multiscale Modeling to Invigorate Engineering Design with Science, Wiley (2012)

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark F-P4

Session: Image and data analysis Image analysis for automated screening and analysis of biological samples using MiniTEM 1

Ida-Maria Sintorn1 Department of Information Technology, University of Uppsala, Uppsala, Sweden

.

E-mail: ida.sintorn@it.uu.se Keywords: Image analysis, machine learning.

Transmission electron microscopy is an important diagnostic tool for analyzing tissue at the nm scale, as well as for detecting and identifying infectious viruses. In this talk I will present image analysis and machine learning approaches and results from ongoing projects with the goals to: a) develop a multi-resolution search and analysis platform for ultrastructural pathologic diagnoses based on the low voltage MiniTEM instrument; b) automatically detect and identify virus particles in nsTEM images.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-I1

Session: Image and Data Analysis Practical Considerations in Image Analysis of Biological Specimens Eija Jokitalo* and Ilya Belevich.

Institute of Biotechnology, Electron Microscopy Unit, P.O. Box 56, 00014 University of Helsinki, Finland. *E-mail: eija.jokitalo@helsinki.fi Keywords: segmentation, quantitative analysis, stereology, large datasets Understanding the structure â&#x20AC;&#x201C; function relationship of cells and cell organelles in their natural context requires multidimensional imaging techniques. As the performance and access to such techniques are improving, the amounts of collected data are growing exponentially posing a question about processing, visualization, and analysis of these large datasets. In conjunction with our 3D-EM and live cell LM projects, we have been developing an open-source software, Microscopy Image Browser (MIB), to effective image processing of multidimensional datasets to improve and ease the full utilization of the acquired data, and to quantitatively analyze morphological features [1-2].

Working in a multiuser environment means that the biological samples that we encounter vary greatly with regard to their origin, shape, size, distribution, and signal intensity, and might require multiple imaging modalities. Therefore, image segmentation workflow and image analysis methods are selected empirically for each project. We also have to take into consideration several practical points before, during and after imaging, which all affect to the analysis. First, the aim of the projects determines the required extent of accuracy in segmentation, which can be lower for visualization of the dataset in comparison to quantitative analysis. Second, segmentation can be done using manual, semi-automatic or automatic segmentation tools, which each have different weaknesses and strengths. Third, for quantitative analysis, we will have to determine countable parameters for a given feature/event, and take into account the requirements of statistical analysis: how the proper sampling is ensured and what is the sufficient number of cells/objects. Finally, we are also testing the possibility to adapt principles of stereology to for the quantitative analysis of big datasets.

[1] I. Belevich, M. Joensuu, D. Kumar, H. Vihinen and E. Jokitalo, PLoS Biology, 14(1): e1002340 (2016).

[2] http://mib.helsinki.fi

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-O1

Session: (Image and Data analysis) Revealing Regions of Correlated Structure in Disordered Carbons

by Scanning Electron Nano Beam Diffraction Espen D. Bøjesen*1, Amelia C. Y. Liu1,2, Timothy C. Petersen2, and Joanne Etheridge.1,3 1Monash

Centre for Electron Microscopy, VIC 3800, Australia.

School of Physics and Astronomy, Monash University, VIC 3800, Australia.

3 Department of Materials Science and Engineering, Monash University, VIC 3800, Australia. *E-mail: espen.bojesen@monash.edu 2

Keywords: Scanning Electron Nanobeam Diffraction, Big Data, Carbon, Amorphous.

The structure of well-ordered carbon allotropes is relatively well understood. However, a similarly reliable structural elucidation of disordered carbon compounds still remains elusive.[1] The correlation length in disordered materials is much shorter than in crystalline materials and standard broad-beam scattering measurements average over too many different local structural environments to specify a unique structure solution for such materials. Thus, critical insights into local symmetries, nano-scale structural heterogeneity and more extended order remain hidden. Scanning electron nanobeam diffraction (SEND) has unique potential to access information across multiple length scales. In a SEND experiment a converged electron probe with dimensions tuned to match the small correlation found in disordered solids is scanned across the sample and an array of spatially resolved electron diffraction patterns is collected. Small signatures of order are revealed in these patterns above the background of structural randomness, providing local symmetry and extended order information.[2] Here we present an exciting new approach to SEND data analysis. It has been tested upon large datasets consisting of 10,000 diffraction patterns collected from 25 x 25nm areas of differently activated carbons, using a Titan 80-300 aberration corrected electron microscope operating at 80kV.The use of principal component analysis and non-negative matrix factorization enables the extraction of subtle features from these large datasets, otherwise hidden upon cursory inspection. The extracted components and loadings are examined to deduce valuable information on the dimensions and orientation of regions of correlated structure,[3] revealing new information about the nature of disordered carbons.

Figure 1: 4D SEND datasets are collected and analysed using a broad range of methods.

[1] Hu, C., Liu, A. C. Y. et al, Carbon, 2015, 85, 119–134.

[2] Liu, A. C. Y., et al., Acta Crystallogr Sect Found Adv, 2015, 71, 473–482.

[3] Cowley, J. M. , Ultramicroscopy, 2002, 90, 197–206.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-O2

Session: Image and Data analysis Refinement Strategy in the Rotation Electron Diffraction Technique Andreas Delimitis*1, Vidar Hansen1, and Jon Gjønnes2 1

Department of Mechanical and Structural Engineering and Materials Science, University of Stavanger, N-4036, Stavanger, Norway 2 Department of Physics, University of Oslo, Gaustadalleen 21, N-0371, Oslo, Norway *E-mail: andreas.delimitis@uis.no Keywords: electron diffraction, rotation method, RED, thermoelectric materials

The applicability of the Rotation Electron Diffraction (RED) method for detailed structure analysis has been broadly demonstrated [1]. Care, however must be taken to account for the remaining dynamical effects in RED, which limits accuracy and precise determination of thermal parameters, such as Debye-Waller factors of thermoelectric materials [2].

In this study, the methodology for incorporating dynamical interactions of the diffracted beams in RED for the determination of structural parameters is presented. Initially, a more precise description and refinement of the diffraction geometry is required. In succession to determining the rotation axis [3], precise calculations and refinement of the incident electron beam direction and beam tilt path will be described, by a combination of Kikuchi line pattern analysis and simulation. HOLZ reflections (as those blue-arrowed in Figure 1 for CoP3) bear a satisfactory kinematical concept and their Kikuchi lines are quite effective for refinement, as they are extremely sensitive to beam tilts and enable calculations with increased precision.

The measurements of excitation errors, sg, for ZOLZ and HOLZ reflections will be also described. It will be shown that these steps are essential for the dynamical approach using Bloch waves. Results of this refinement strategy in the study of thermoelectric materials will be presented and evaluated.

Acknowledgements: The authors wish to express their gratitude to Mika Buxhuku and Peter Oleynikov for their contributions to this work.

Figure 1: (a) Experimental and (b) simulated CoP3 patterns, beam tilted at 0.75o from [3 1 7].

[1] D. Zhang, P. Oleynikov, S. Hovmöller and X. Zou, Z. Kristallogr 225, 94–102 (2010).

[2] M. Buxhuku, V. Hansen and J. Gjønnes, Micron 101, 103-107 (2017).

[3] M. Buxhuku, V. Hansen, P. Oleynikov and J. Gjønnes, Microsc. Microanal. 19, 12761280 (2013).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-O3

Session: Image and Data analysis Bayesian Analysis of Noisy Scanning Transmission Electron Microscopy Images for Single Atom Detection J. Fatermans*1,2, A. J. den Dekker2,3, K. MĂźller-Caspary1, I. Lobato1, and S. Van Aert1.

1

2

EMAT, University of Antwerp, Groenenborgerlaan 171, 2020 Antwerp, Belgium.

Imec-Vision Lab, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.

3 DCSC, Delft University of Technology, 2628 CD Delft, The Netherlands. *E-mail: Jarmo.Fatermans@uantwerpen.be

Keywords: STEM, single atom detection, parameter estimation, model-order selection.

In principle, scanning transmission electron microscopy (STEM) is a powerful technique to image single atoms. However, to avoid beam damage, the incoming electron dose should be kept low enough resulting into images exhibiting a low signal-to-noise ratio (SNR). In general, visual inspection of such images leads to biased structure information. To overcome this problem, the maximum a posteriori (MAP) probability rule is proposed, which, by combining parameter estimation and model-order selection, enables one to detect single atoms with high reliability.

The validity and usefulness of the MAP probability rule has been demonstrated for experimental and simulated STEM images of samples of arbitrary shape, size and atom type.

First, the most probable structure indicated by the MAP probability rule from an experimental low SNR image of SrTiO3 (figure 1(a)) corresponds to the expected crystal structure of SrTiO3 in [100] direction (figure 1(b)). Next, individual atoms near the edge of a Au nanorod (figure 1(c)) [1] have been successfully detected by the MAP probability rule (figure 1(d)).

Moreover, for lower incoming electron dose (figure 1(e)) the MAP probability rule obtains reliable results. Only one extra atom has been detected where the image did not include an atom, indicated by the arrow in figure 1(f). Finally, the most probable structure from an experimental image of a Ge cluster (figure 1(g)) [2] is shown in figure 1(h).

Figure 1: (a), (c), (e), (g) Experimental high-angle annular dark-field (HAADF) STEM image of SrTiO3 [100], experimental HAADF STEM image of a Au nanorod, simulated HAADF STEM image of a Au nanorod, and experimental HAADF STEM image of a Ge cluster, respectively. (b), (d), (f), (h) Most probable parametric models from (a), (c), (e) and (g), respectively, with the detected atom columns shown in red.

[1] S. Van Aert et al., Physical Review B 87, 064107 (2013).

[2] S. Bals et al., Nature Communications 3, 897 (2012).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-O4

Session: Image and data analysis Point of Origin? Application of Automated Mineralogy in Archaeology Kirsten I. Kling*1, Agnieszka M. Bystron2, and Theis I. Sølling3.

1

Centre for Electron Nanoscopy (CEN), Technical University of Denmark, Fysikvej 307, 2800 Kgs. Lyngby, Denmark.

2 Department of Cross-Cultural and Regional Studies, University of Copenhagen, Karen Blixens Plads 8, 2300 Copenhagen.

3 Department of Chemistry, University of Copenhagen, Universitetsparken 5, 2100 Copenhagen, Denmark. *E-mail: kili@dtu.dk Keywords: QEMSCAN, automated SEM, mineralogy, archaeology.

Al Zubarah in North-Western Qatar was a rich regional center for trade prior to its abandonment in the late 19th century [1]. Regardless of this, not much information exists in terms of recorded trade routes and partners. Applying automated mineralogy in a Scanning Electron Microscope (SEM), we attempt to determine the origin of pottery sherds excavated from Al Zubarah by mapping the mineralogical composition and texture of small fragment samples. The yet obtained results are not entirely conclusive, but they do show that automated mineralogy in form of the here presented QEMSCAN process is a highly valuable method and applicable to fundamental question in archaeology.

Quantitative Evaluation of Mineralogy is anchored around a state-of-the-art SEM instrument (FEI QEMSCAN), equipped with tw o energy dispersive X-ray (EDS), secondary electron (SE) and backscattered electron (BSE) detectors. Using automated stage driving and programmed processes, large areas on multiple samples are scanned and data is obtained with high spatial resolution.

Storage Jar, Oxidised Coarse Ware Storage Jar, Oxidised Coarse Ware

1 cm

Figure 1: Preparation of small fragment samples reveals a diverse mineralogy of the sherd.

[1] Richter, Tobias, Faisal Abdulla al-Naimi, Lisa Yeomans, Michael House, Tom Collie, Pernille Bangsgaard Jensen, Sandra Rosendahl, Paul Wordsworth, and Alan Walmsley. "The 2010-2011 excavation season at al-ZubÄ rah, north-west Qatar (poster)." In Proceedings of the Seminar for Arabian Studies, pp. 331-339. Archaeopress, 2012.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-O5

Session: Image and Data Analysis Deformable Curves for Outlining Objects Directly From Projections Vedrana Andersen Dahl*, Jakeoung Koo, Per Christian Hansen and Anders Bjorholm Dahl.

Technical University of Denmark, Department of Applied Mathematics and Computers Science, Richard Petersens Plads, DK-2800 Lyngby *E-mail: vand@dtu.dk Keywords: tomographic reconstruction, deformable models, segmentation, meshing.

Processing of X-ray tomographic projection data usually starts by computing a reconstructed 2D or 3D image, often followed by an image segmentation step. Each of these steps introduces errors and artefacts, which is especially pronounced when data is noisy or incomplete. This has motivated the development of methods that combine the reconstruction and the segmentation step. When the object under study consists of a number of domains with approximately homogeneous absorption coefficients, the segmentation may be obtained by evolving the curve in the reconstruction domain.

We developed an efficient algorithm that computes a segmented reconstruction directly from X-ray projection data [1]. Our algorithm uses a parametric curve to define the segmentation. Unlike similar approaches, which are based on level-sets, our method avoids a pixel or voxel grid; hence the number of unknowns is reduced to the point-set outlining the curve, and the attenuation coefficients of the segments. Trough systematic tests on synthetic data, as illustrated in Fig. 1, we demonstrated a high robustness to the noise, and an excellent performance under a small number of projections. Recently we tested our method on the real tomography data, yielding promising results shown in Fig. 2.

Figure 1: A synthetic test object, ground truth sinogram consisting of 15 projection angles and 200 detector pixels, a sinogram corrupted by Gaussian noise (noise level 0.1), and an illustration of a curve evolution obtained when reconstructing the noisy sinogram.

Figure 2: A photograph of a small plastic object, a single image from a sequence of the Xray projections, a sinogram corresponding to a one slice through the object and a reconstructed object outline represented by 200 points.

[1] V.A. Dahl, A.B. Dahl and P.C. Hansen. "Computing segmentations directly from X-ray projection data via parametric deformable curves." Measurement Science and Technology 29.1 (2017): 014003.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-O6

Session: Image and data analysis Large Area Crystal Phase Mapping by Scanning Electron Diffraction and Machine Learning Data Analysis Julie S. Nilsen*1, Håkon W. Ånes2, Ingrid M. Andersen1, Dingding Ren3, Helge Weman3, Bjørn-Ove Fimland3, Antonius T.J. van Helvoort1.

1

Department of Physics, 2Department of Material Science and Engineering, 3Department of Electronic Systems, Norwegian University of Science and Technology (NTNU), NO-7491, Trondheim, Norway. *E-mail: julie.s.nilsen@ntnu.no Keywords: Precession electron diffraction, non-negative matrix factorization, nanowires.

Modern microscopy systems allow automatic collection of large data sets, which require novel data analysis strategies. Here we present crystal phase mapping based on scanning precession electron diffraction (SPED) and non-negative matrix factorization (NMF) data evaluation. The results are compared to conventional transmission electron microscope (TEM) techniques on the same area. As an example we use 400 nm thick GaAs nanowires (NWs) with six axial GaAsSb superlattices showing room temp. single-mode lasing in the near-IR range [1]. Data analysis was done in the open-source Python library HyperSpy [2].

Conventional TEM (selected-area electron diffraction (SAED), high-resolution (HR) and dark-field (DF) imaging) shows that each superlattice consists of ten zinc blende segments in two orientations (ZB1, ZB2) with spacers of mixed phase, and wurtzite (WZ) segments at the ends (Fig. (a-c)). However, this work is laborious and spatially restricted. With SPED, an electron diffraction pattern is collected for each precessed probe position (here 73 910) from which the crystal phase can be determined and virtual dark-field (VDF) images can be constructed (Fig. (d-g)). However, thickness effects (Fig. (h-i)) and the data set size make automatic phase recognition challenging. Alternatively, NMF analysis can be applied [3]. Six NMF components are required to extract the three expected phases (Fig. (j-k)), but using more components can give additional information. Spatial resolution, the effect of specimen thickness and bending will be discussed. It will be shown that this approach can analyze large areas (5 µm2) with nm-size resolution, well beyond what is achievable by conventional TEM.

Figure: Comparison of crystal phase mapping of a NW with conventional TEM (a-c), SPED (d-i) and NMF analysis of SPED data (j-k). Red: ZB1, Green: ZB2, blue: WZ. [1] D. Ren et al., Nano Lett., Article ASAP. DOI: 10.1021/acs.nanolett.7b05015 (2018).

[2] F de la Peña et al., Hyperspy 1.3 Zenodo. https://doi.org/10.5281/zenodo.583693, (2017).

[3] A. Eggmann et al., Nature Comm. 6, 7267 (2015).

Acknowledgements: RCN for the support of NORTEM (197405) & NANO2021 (239206).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-P1

Session: Image and Data Analysis The Morphology Affects Bacterial Virulence in Mycobacteria Nagatoshi Fujiwara*1, Minoru Ayata2, Hiroyuki Yamada3, Takashi Naka1, Hirotaka Kuwata4, and Shinji Maeda5, 1

2

Tezukayama University, Nara 631-8585, Japan Osaka City University Graduate School of Medicine, Osaka 545-8585, Japan 3 Research Institute of Tuberculosis, JATA, Tokyo 204-8533, Japan 4 Showa University School of Dentistry, Tokyo 142-8555, Japan 5 Hokkaido University of Science, Hokkaido 006-8590, Japan *E-mail: fujiwara@tezukayama-u.ac.jp

Keywords: glycopeptidolipid, phenolglycolipid, Mycobacterium smegmatis, Mycobacterium bovis BCG Mycobacterium smegmatis is a rapidly growing, nonpathogenic mycobacterium, and has been used as a tool for molecular analysis of mycobacteria. The M. smegmatis mc2155 strain was a smooth colony, and the M. smegmatis J15cs strain was a rough and dry colony. The SEM analysis clarified the difference of the bacterial morphology in these two strains. It is caused that the J15cs strain has deleted apolar glycopeptidolipids (GPLs) in the cell wall, and affects the bacterial size, morphology, and survival in host cells [1, 2]. The similar phenomenon was observed with Mycobacterium abscessus strains [3]. On the other hand, Mycobacterium bovis BCG Tokyo 172 substrain is a predominant World Health Organization (WHO) Reference Reagent for the BCG vaccine. The BCG Tokyo 172 substrain was reported to consist of two subpopulations with different colony morphologies, smooth and rough. The smooth colony had a characteristic 22 bp deletion in Rv3405c of the RD16 region (type I), and the rough colony was complete in this region (type II). We determined the lipid compositions and biosynthesis of type I and II, to check the relationship between the morphological difference and lipid phenotypes. The SEM analysis showed that the cell size of type I was 1.5 times longer than that of type II. Phenolglycolipid (PGL) and phthiocerol dimycocerosate (PDIM) were expressed only in type I. We found that the existence of PGL/PDIM in types I and II is caused by a mutation of ppsA gene responsible for PGL/PDIM biosynthesis. PGL suppressed the host recognition of total lipids via Toll-like receptor 2, and this suggests that PGL is antigenic and involved in host responses acting as a cell wall component [4].

In conclusion, the existence of lipid components in cell wall affected bacterial morphologies, host-pathogen interaction, and bacterial virulence.

Figure: The morphologies of Mycobacterium species.

[1] N. Fujiwara et al., Tuberculosis (Edinb). 92, 187-192 (2012).

[2] N. Fujiwara, et al., PLoS One 10(5):e0126813 (2015).

[3] J. Whang et al., Cell Death Dis. 8(8):e3012 (2017).

[4] T. Naka et al., J. Biol. Chem. 286(51), 44153-44161 (2011).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-P2

Session: Image and Data analysis Automated Image and Analysis of Pharmaceutical Particles Using a Tabletop Low Voltage TEM Lin Zhu*1, Martin Ryner1, Gustaf Kylberg1 and Ida-Maria Sintorn1,2 1

2

Vironova AB, Gävlegatan 22, 11330 Stockholm, Sweden.

Uppsala University, Department. of Information Technology, 75105 Uppsala, Sweden. *E-mail: johan.harmark@vironova.com Keywords: Instrument, Automation, Image Analysis.

The development of nanoparticle technologies has been significantly increasing over the past decade, with applications in a broad scope of fields, as well in material sciences as in life sciences, and the demand on particle characterization has been growing accordingly. Amongst the methods available for particle characterization, Electron Microscopic technologies in contrast to indirect methods like Dynamic Light Scattering and Nano Tracking Analysis result in high resolution images that provide morphological information and unambiguous particle identification. EM however requires an experienced operator and can be time and resource consuming. A method allowing to simplify the analysis process with an automated image acquisition of samples as well as an automatic image analysis on a low-voltage electron microscope is proposed herein.

Such automation capabilities prove relevant for example in the domain of the vaccine industry, where new requirements from the United States Pharmacopeial Convention (USP) and the U.S. Food and Drug Administration (FDA) put pressure for root cause analysis to understand impact on changes during process and product development. The new demands involve morphological characterization of the particles present in the formulations, typically nanoparticles like viruses, virus-like particles and adjuvants in the range of 0.01 - 0.1 Âľm. When entering production phases of such products, the high throughput of specimen to be analyzed can be a limiting factor for electron microscopy to be used as a method of choice. We therefore introduced a process that can allow a rapid and efficient characterization of specimen, using a low voltage instrument, the MiniTEMâ&#x201E;˘.

The instrument consists of a low voltage (25kV) transmission electron microscope and software including a simple control of the microscope s and automated features for imaging and analysis.

The instrument is small and requires only a single standard power outlet, allowing the system to be placed in any lab or office area. The simplified design of the instrument makes its use easier to apprehend for users with limited experience in electron microscopy.

Detected particles within a preset size range are, while screening, automatically measured and characterized based on assigned attributes for each type of particle. If desired the screening can be set to stop when sufficient information has been acquired. Stop criteria such as a minimum number of detected particles or statistical stability of the size distribution can be used.

A case study involving the characterization of biological nanoparticles in which the specimen was automatically analyzed will be presented and discussed.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-P3

Session: Image and Data analysis Quantifying Aerosol Size, Shape, and Composition Distributions via Impaction and Automated Electron Microscopy Analysis A. Brostrøm*1,2, K. Mølhave1, and K. I. Kling1.

1

2

Technical University of Denmark, Kgs. Lyngby, Denmark National Research Centre for the Working Environment, Copenhagen, Denmark. *E-mail: abbl@dtu.dk

Keywords: Aerosol, Particle Characterization, Impaction, Scanning Electron Microscopy.

Air pollution is one of the major contributors to the global burden of disease, with particulate matter (PM) as one of its central concerns [1]. However, current standard measurement techniques bring no knowledge of particle composition or shape, which has been identified as crucial parameters in toxicological studies [2]. Accordingly, additional measurement techniques are needed to provide a more detailed characterization and to establish standard procedures for measuring and regulating PM pollution.

Automated Scanning Electron Microscopy (SEM) coupled with Energy Dispersive X-ray Spectroscopy (EDS) can provide both detailed physical and elemental composition single particle information. It can furthermore obtain sufficient data for statistical analysis by systematically mapping large areas of a sample without user intervention. As a result the technique can provide size and morphology resolved elemental composition data of aerosol populations. However, the technique lacks standard procedures for collecting and imaging particles, ensuring a representative and reproducible sample description.

Here we present the current development of a standard operating procedure for sampling aerosols via impaction [3] directly onto TEM grids, for automated SEM/EDS analysis. The method was tested by collecting an aerosol of 40, 100, 200, and 500 nm spherical latex beads (PSL). From the deposition pattern on the TEM grid an imaging routine was established to obtain a representative particle size distribution (PSD), where a series of images was acquired from one edge of the impact area to the other, going through its center. To verify the imaging routine, the average PSD obtained from the series of images was compared to the PSD measured via Scanning Mobility Particle Sizer (SMPS), which was corrected for impactor collection efficiency (Ceff), shown in Figure 1. Good agreement was found between the two PSDs, showing that SEM analysis can be used to obtain representative PSDs of impacted aerosols. This enables the use of SEM/EDS to give detailed descriptions of collected aerosols e.g. size, aspect ratio, and circularity as well as elemental composition of individual particles.

Figure 1: PSD of 40, 100, 200, and 500 nm PSL aerosol measured by automated SEM analysis (blue) and SMPS (red). Impactor cut-off marked as D50.

[1] P. J. Landrigan et al., The Lancet Commission on pollution and health (2017).

[2] G. Oberdorster et al., Environmental Health Perspectives, 113(7), 823–839 (2005).

[3] K. Kandler et al., Atmospheric Environment, 41(37), 8058–8074 (2007).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-P4

Session: Image and Data Analysis Understanding UD Fibre-reinforced Polymers through X-ray Imaging and Individual Fibre Tracking Monica J. Emerson*1, Anders B. Dahl1, Vedrana A. Dahl1, Knut Conradsen1, Ying Wang2, Philip J. Withers2, Kristine M. Jespersen3 and Lars P. Mikkelsen4.

1

Image Analysis and Computer Graphics, DTU Compute, Kgs. Lyngby, Denmark.

2 Henry Moseley X-ray Imaging Facility, School of Materials, UoM, UK. 3 Kanagawa Institute of Industrial Science and Technology, Waseda University, Tokyo, Japan.

4 Composite and Materials Mechanics, DTU Wind Energy, Roskilde, Denmark. *E-mail: monj@dtu.dk Keywords: dictionary-based image segmentation, geometrical characterisation, nondestructive testing, fibre composites, micro-computed tomography X-ray computed tomography (CT) is a powerful tool for characterising materials for its ability to reveal their internal structure in a non-destructive manner. The recent advances in Xray imaging have brought high-resolution X-rays to laboratory sources, making this tool available to a broader public. Additionally, thanks to the developments in ultra-fast X-ray imaging at synchrotron beamlines, it is now possible to capture the very fast structural changes inside materials under realistic working conditions, e.g. in operation or under loading.

There is a need for advanced image analysis methods that can exploit the information contained in these 3D and 4D data-sets of high spatial and temporal resolution, which often contain image artefacts and noise. We have developed a method to characterise the geometry of materials reinforced with long fibres [1], such as glass and carbon fibre reinforced polymers.

The method is based on segmenting individual fibres and the task is specially challenging when the image is noisy and its resolution is limited, because the fibres are densely packed. A limited spatial resolution might arise from the need of performing fast scans, to capture the sudden micro-structural changes that happen when reaching the compositeâ&#x20AC;&#x2122;s collapse load, and will facilitate scanning large fields of view containing many fibres, necessary to ensure representative characterisations of a materialâ&#x20AC;&#x2122;s micro-structure.

Due to the robustness of our method to image quality [2], we have been able to characterise fibre orientations and diameter distributions in complete bundles, relevant for investigating the effect of the design and manufacturing processes on the mechanical properties of the materials.

Moreover, we have applied our methodology to study the behaviour of a fibre composite under compressive loading. Following the changes in each individual fibre under progressive loading conditions, and correlating these with the initial structure of the material, can reveal the precursors to the very complex damage mechanisms that affect fibre composites.

Figure 1: Characterisation of the micro-structure inside UD fibre reinforced composites.

[1] M. J. Emerson, K. M. Jespersen, A. B. Dahl, K. Conradsen, L. P. Mikkelsen, Individual fibre segmentation from 3D X-ray computed tomography for characterising the fibre orientation in unidirectional composite materials. Composites Part A 97 (2017), 83-92.

[2] M. J. Emerson, V.A. Dahl, K. Conradsen, L. P. Mikkelsen, A. B. Dahl, Statistical validation of individual fibre segmentation from tomograms and microscopy, Compos. Sci. and Technol. 160 (2018) 208-215.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-P5

Session: Image and data analysis Can we train a single deep learning model to detect and segment nuclei images acquired with any microscope or staining modality?

ASM Shihavuddin*1, Florian Gawrilowicz2, Niels Jeppesen3, Rasmus Reinhold Paulsen4.

1

Postdoctoral researcher, DTU compute, Technological University of Denmark, Lyngby 2 PhD student, DTU compute, Technological University of Denmark, Lyngby 3 PhD student, DTU compute, Technological University of Denmark, Lyngby 4 Associate professor, DTU compute, Technological University of Denmark, Lyngby *E-mail: shihav@dtu.dk

Keywords: nuclei segmentation, deep learning, Faster RCNN, Inception resnet v2, segnet Human body comprises around 30 trillion cells each containing nucleus full of DNA that are studied regularly for research in drug discovery, understanding body functionality, etc. Lack of automated and reliable segmentation of the cell nuclei is one the major bottleneck in reaching the potential speed of growth in these fields of science. Over the years many algorithms been proposed for nuclei segmentation [1], however most of them perform well only for specific cell, staining or imaging type [2]. In this work, we developed a framework with deep learning methods that can accumulate knowledge about nuclei from expert biologists through annotations and convert it into a tool for nuclei segmentation. In the presented work, we first trained an Inception-resnetV2 [2] network with Faster R-CNN detection framework to initially detect the nucleus in terms of bounding box. In the second part, we trained a segmentation network with VGG16 architecture to segment the nucleus within the bounding box. The first stage was trained on 670 images and the second stage was trained on 32000 nuclei. On the test sets of 65 images, we recorded .5 mean average precision (map), when averaged over 0.5 to 0.95 (with 0.05 interval) intersection by overlap (IoU) ratio of segmentation with ground truth. Figure 1 illustrates the nuclei segmentation results on some of the challenging examples in the test set. The resulted accuracy shows the model is able to learn about the required visual properties for identifying nuclei and execute it on unseen images with reliable accuracy.

Figure 1: Nuclei segmentation result on input images of varies imaging modalities A, C. E, G & I are illustrated (with jet color map) in B, D, F, H & J respectively.

[1] Xing, F. and Yang, L., 2016. Robust nucleus/cell detection and segmentation in digital pathology and microscopy images: a comprehensive review. IEEE reviews in biomedical engineering, 9, pp.234-263.

[2] Meijering, E., 2012. Cell segmentation: 50 years down the road [life sciences]. IEEE Signal Processing Magazine, 29(5), pp.140-145.

[3] Szegedy, C., Ioffe, S., Vanhoucke, V. and Alemi, A.A., 2017, February. Inception-v4, inception-resnet and the impact of residual connections on learning. In AAAI (Vol. 4, p. 12).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-P6

Session: Image and Data analysis Macrophage-uptake of sialic acid-targeted molecularly imprinted polymers (MIPs) Louise Sternbæk*1, Sudhirkumar Shinde2, Börje Sellergren2, Kersti Alm1 and Anette Gjörloff Wingren2.

1

2

Phase Holographic Imaging,Scheelevägen 22, 223 63 Lund, Sweden.

Biomedical Science, Health and Society, Malmö University, Malmö, Sweden. *E-mail: louise.sternbaek@phiab.se

Keywords: cancer, molecular imprinted polymers, fluorescence microscopy, flow cytometry, digital holographic microscopy.

Sialic acid (SA) is a cell surface glycan, which has a decisive role in many cell activities including differentiation, proliferation, and the immune response [1]. The amount of SA has been found to correlate with cancer, with an upregulation on more aggressive cancers.

Therefore, there is a great interest in developing methods for detection of SA on cancer cells.

We are screening SA on cancer cell lines by using fluorescent molecularly imprinted polymers, SA-MIPs [2, 3].

Here we aim to investigate the possible uptake of SA-MIPs by macrophages in culture. The uptake will be analysed by flow cytometry, fluorescence microscopy and digital holographic microscopy. We also aim to investigate the viability of cells after ingestion of SA-MIPs, and whether the uptake is SA-specific.

Future aims are to use MIPs as a diagnostic tool in vivo. These results will give an understanding of the possible uptake by cells of the immune system in the body.

Figure 1: Fluorescence microscopy showing the cells have internalized the molecular imprinted polymers after 24 hours incubation.

[1] Varki, A. et al. (2009). Essentials of Glycobiology, 2nd edition, Chapter 14, Sailic Acids.

[2] Shinde, S. et al. (2015). Sialic Acid-Imprinted Fluorescent Core-Shell Particles for Selective Labeling of Cell Surface Glycans. Journal of the American Chemical Society, 20:13:02.

[3] El-Schich, Z. et al. (2016). Different expression levels of glycans on leukemic cells – a novel screening method with molecular imprinted polymers (MIP) targeting sialic acid. Tumour Biology, 37:10

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark G-P7

Session: Imaging multicellular systems, Live imaging of single cells, Correlative Light and Electron Microscopy (CLEM) Big data approaches for computational phenotyping Thomas Walter1, 2, 3 1

MINES ParisTech, PSL-Research University, CBIO-Centre for Computational Biology, 35 rue St Honoré 77300 Fontainebleau, France 2 Institut Curie, 75248 Paris Cedex ,France.

3 INSERM U900, 75248 Paris Cedex, France. *E-mail: Thomas.Walter@mines-paristech.fr Keywords: machine learning, artificial intelligence, bioimage informatics, Spatial Transcriptomics, High Content Screening, histopathology.

In the field of bioimaging, we dispose of the technological tools to perform imaging experiments at an unprecedented scale and thus to generate extremely large and complex image data sets, that can be readily qualified as “big data”. For instance, in High Content Screening (HCS), a large number – typically tens or hundreds of thousands - of different experimental conditions can be tested with respect to their effect on cells and organisms, as measured by microscopy. Examples include genetic screens aiming at inferring gene function from their lossof-function phenotypes and drug screens, where drugs are characterized by their phenotypic effect. But the big data revolution in Bioimaging is not limited to HCS. Another example is histopathology, where it is not the number of experiments, but the complexity of each single image that contributes to the big data aspect. Indeed, a single stained tumor section typically contains hundreds of thousands of cells. All cells can be phenotyped, and their spatial distribution can be assessed and analyzed, which is a formidable challenge.

Here, I will present the methodological framework that allows us to exploit such large and systematic data sets. First, I will review the techniques we and others have developed in order to infer gene function from live-cell imaging data in the context of loss-of-function screens [1], [2], [3]. Second, I will focus on the emerging field of spatial transcriptomics. Gene expression has been studied for many years using microarrays and RNAseq. Today, we can study the spatial aspects of gene expression at a large scale with scalable single molecule FISH techniques [4]. These recently developed imaging methods also come with interesting new questions for the computational analysis: single RNAs can be automatically detected and their spatial distribution analyzed with newly designed features and machine learning methods. Importantly, there is little prior knowledge available for this type of data. We have thus built a virtual environment in order to simulate RNA localization patterns inside cells which we can use as benchmark for validation and method refinement. Third, I will present new approaches to analyze and computationally phenotype cells in their tissular context for histopathology. One of the major problems in computational approaches for this data is the segmentation of nuclei, which is notoriously difficult given the variability of the data. Here, I present a new technique for nuclei segmentation based on a combination of deep learning with more traditional methods [5]. This method will allow us to analyze large cohorts of patient data with respect to their cellular and tissular phenotypes and to relate these descriptors to clinical variables.

[1] B. Neumann, T. Walter et al., Nature 464 (7289), 721–7 (2010).

[2] M. Mall et al., Journal of Cell Biology 198(6), 981-990, (2012).

[3] A. Schoenauer Sebag et al., Bioinformatics, 31 (12), i320-i328 (2015).

[4] N. Tsanov et al., Nucleic acids research, 44 (22), e165-e165 (2016).

[5] P. Naylor et al., Int. Symposium on Biomed. Imaging (ISBI), 933–36 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-I1

Session: Imaging multicellular systems, Live imaging of single cells, Correlative Light and Electron Microscopy (CLEM) Quantitative Imaging of Intestinal Organoid Development Gustavo de Medeiros1, Denise Serra1,2, Urs Mayr1,2, Ilya Lukonin1,2, Andrea Boni3, Katrin Volkmann1, Michael Stadler1, Petr Strnad3, Panagiotis Papasaikas1, Annick Waldt3, Guglielmo Roma3, and Prisca Liberali*1,2.

1

Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, 4058 Basel.

2 University of Basel. Petersplatz 1, 4001 Basel.

3 École polytechnique fédérale de Lausanne (EPFL), School of Life Sciences, Station 19 1015 Lausanne. *E-mail: prisca.liberali@fmi.ch Keywords: intestinal organoids, high-content screening, light-sheet microscopy.

The development of intestinal organoids follows a self-organized, de novo morphogenesis through the emergence of an asymmetrical system from an initially homogeneous population of cells [1], ultimately leading to fully-grown organoids containing most of the functional properties of the intestine [1,2].

Our goal is to quantitatively understand the molecular and mechanical cues driving the development of intestinal organoids. To understand this emergent self-organizing system we need to record how intestinal organoids change morphologically and genetically, in space and time [3]. To overcome these challenges the laboratory focuses on developing new image-based technologies that can provide multivariate features for numerous organoids and long term dynamical data.

The high-content image-based screening pipeline we have developed allows us to extract multivariate feature sets from fixed time-points of organoids cultured during 5 days. With the extracted features set we infer growth trajectories in the multivariate feature space, making it possible to quantitatively describe the development of different phenotypes.

Furthermore, organoid development from single stem cells can be recorded with highspatiotemporal resolution with our dedicated light-sheet microscope for organoid imaging [4].

With this technology we can focus on recording the driving morphogenetic dynamics that ultimately lead to a fully asymmetrical, fully functional organoid. Furthermore, this system also allows precise optical manipulations to be performed live, allowing us to perturb the system as it develops.

The combination of multivariate time-courses with light-sheet time-lapse imaging allows us to understand how single cells have the intrinsic ability to generate emergent, self-organized structures such as organoids.

[1] T. Sato et al., Nature. 459, 262–265 (2009).

[2] A. E. Shyer, T. R. Huycke, C. Lee, L. Mahadevan, C. J. Tabin, Cell. 161, 569–580 (2015).

[3] A. C. Rios, H. Clevers, Imaging organoids: a bright future ahead. Nature Methods (2018).

[4] P. Strnad et al., Nature Methods. 13, 139 (2015).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-I2

Session: Live imaging of single cells Characterizing Cancer Stem Cell Movement and Division using Digital Holographic Imaging in Combination With Fluorescence Sofia Kamlund*1,2, Xiaoli Huang1, Birgit Janicke2, Kersti Alm2, and Stina Oredssson1.

1

Department of Biology, Lund University, Sölvegatan 35, 223 62 Lund, Sweden.

2 Phase Holographic Imaging AB, Scheelevägen 22, 223 63 Lund, Sweden. *E-mail: sofia.kamlund@biol.lu.se

Keywords: digital holography, cancer stem cells, longitudinal tracking, time-lapse imaging, fluorescence.

While the heterogeneity of tumors has been known for a while, many assays used for investigating cancer cells are still population-based. Those assays are often fast, however lacking the ability to distinguish differences in behavior between cells within a population.

Increasing evidence indicate that a small population of cells in tumors, called cancer stem cells, are responsible for inducing tumor growth and metastasis [1]. The demand for cancer stem cell targeting drugs, as well as more knowledge about cancer stem cells is increasing.

We have previously shown that longitudinal tracking of human breast cancer JIMT-1 cells, using digital holography, can distinguish between cells with different kinds of behavior within the cell population and that treatment with salinomycin changes the behavior of the cells [2].

Salinomycin is an antibiotic ionophore that targets cancer stem cells [3]. Salinomycin has previously been shown to inhibit cell proliferation based on population-derived data [4].

However, our study of individual cells shows that even after 72 hours of salinomycin treatment, there remains a small population of dividing cells that is not detected in populationbased data. The same study also shows that cell migration is affected by salinomycin treatment [2]. However, in that study we could not connect effects on cell proliferation and migration behavior with the phenotype of the cells. In breast cancer, the cancer stem cell population has been identified as CD44+ESA+CD24-/low [1]. Therefore, we have separated JIMT-1 cells into sub-populations using magnetic bead separation, based on the expression of CD24 and ESA (CD24+, CD24-ESA+, CD24-ESA-). We have analyzed the isolated subpopulations using flow cytometry and seeded them and tracked them for 72 hours using digital holography. Furthermore, we have developed a new combinational assay of digital holography and fluorescence microscopy where we can distinguish between different cell populations based on CD24 expression. The different sub-populations were found to proliferate at different rates and had different migration behavior. Thus, using this assay, it is possible to follow drug treatment of different sub-groups of a cell population.

[1] [2] [3] [4]

M. Al-hajj, M. S. Wicha, A. Benito-hernandez, S. J. Morrison, and M. F. Clarke, PNAS 100, 3983–3988 (2003).

S. Kamlund, D. Strand, B. Janicke, K. Alm, and S. Oredsson, Cell Cycle 4101, 1–11 (2017).

P. B. Gupta, T. T. Onder, G. Jiang, K. Tao, C. Kuperwasser, and R. A. Weinberg, Cell 138, 645–659 (2009).

X. Huang, B. Borgström, S. Kempengren, L. Persson, C. Hegardt, and D. Strand, BMC Cancer, 16, 1–13 (2016).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-O1

Session: Correlative Light and Electron Microscopy (CLEM) How To Characterize Individual Nano-Size Liposomes With Simple Self-Calibrating Fluorescence Microscopy Kim I. Mortensen*1, Chiara Tassone1, Nicky Ehrlich1, Thomas L. Andresen1, and Henrik Flyvbjerg1 1

Department of Micro- and Nanotechnology, Technical University of Denmark, Kongens Lyngby, DK-2800, Denmark. *E-mail: kim.mortensen@nanotech.dtu.dk

Keywords: liposomes, vesicles, single-particle analysis, dual-color fluorescence microscopy, lamellarity.

Nano-size lipid vesicles are used extensively at the interface between nanotechnology and biology, e.g. as containers for chemical reactions at minute concentrations and vehicles for targeted delivery of pharmaceuticals. Typically, vesicle samples are heterogeneous as regards vesicle size and structural properties (Fig. 1). Consequently, vesicles must be characterized individually to ensure correct interpretation of experimental results. Here [1] we do that using dual-color fluorescence labeling of vesiclesâ&#x20AC;&#x201D;of their lipid bilayers and lumens, respectively (Fig. 1). A vesicle then images as two spots, one in each color channel. A simple image analysis determines the total intensity and width of each spot. These four data all depend on the vesicle radius in a simple manner for vesicles that are spherical, unilamellar, and optimal encapsulators of molecular cargo. This permits identification of such ideal vesicles. They in turn enable calibration of the dual-color fluorescence microscopy images they appear in. Since this calibration is not a separate experiment but an analysis of images of vesicles to be characterized, it eliminates the potential source of error that a separate calibration experiment would have been. Non-ideal vesicles in the same images were characterized by how their four data violate the calibrated relationship established for ideal vesicles. In this way, our method yields size, shape, lamellarity, and encapsulation efficiency of each imaged vesicle. Applying this procedure to extruded samples of vesicles, we found that, contrary to common assumptions, only a fraction of vesicles are ideal.

Figure 1: Fluorescence microscopy allows characterization of individual, dual-color labeled (red, lipid bilayer; blue, lumen) vesicles with regards to size, shape, lamellarity, and encapsulation efficiency.

[1] K.I. Mortensen, C. Tassone, N. Ehrlich, T.L. Andresen, and H. Flyvbjerg, Nano Letters (2018); doi: 10.1021/acs.nanolett.7b05312

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-O2

Session: Imaging multicellular systems or Live imaging of single cells How Nanoparticles Disturb The Lipid Bilayer Vesicles Meriem Er-Rafik*,1, Khalid Ferji 2, Olivier Sandre 2, Carlos M. Marques 1, Jean-Francois Le Meins 2, Marc Schmutz 1 1

Institut Charles Sadron, Université de Strasbourg-CNRS UPR 22, 67034 Strasbourg, France Laboratoire de Chimie des Polymères Organiques (LCPO), Université de Bordeaux, CNRS, 33607 Pessac, France *E-mail: meriem.er-rafik@ics-cnrs.unistra.fr

2

Keywords: nanoparticle, liposome, leaflet bilayer membrane.

Innumerous molecules and particles, from simply water to complex proteins or selfassembled small liposomal carriers, can frequently interact with the cell membrane with possible modification on it. Despite a constant increase of the variety of new particles or molecular assemblies, due to rapid progress in nanotechnology, the molecular features determining how is the membrane behaviour with respect to a given molecule are not yet elucidated.

Here, we will present a new mechanism of the lipid bilayer of liposomes behaviour in presence of nanoparticles investigated by cryo-electron tomography. Cryo-electron microscopy is a relevant technique allowing not only to inspect the structure of the membrane, by resolving for instance the two leaflets of the bilayer, but reveals also geometric features of nanoparticles such as size and shape that play an important role for the potential interaction with lipid bilayer (fig. 1). Cryo-electron tomography resolve in 2D and 3D space the relative positions of particles and membranes, providing insight into the interplay between particlelipid interactions and the ensuing bilayer transformations.

Figure 1: Silica nanoparticles – DOPC liposome interaction after 20 sec.

We wish to thank for the financial support the research association ANR (Agence Nationale de Recherche) of the project ANR-12-BS08-0018-01 and the French society of microscopy (SFµ).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-O3

Session: Single cell imaging Quantitative Image Based Cytometry For Cell Biology Research Luis Toledo*1 Center for Chromosome Stability University of Copenhagen, Blegdamsvej 3B, 2200 N. Copenhagen, Denmark. *E-mail: ltoledo@sund.ku.dk 1

Keywords: quantitative imaging, single cell imaging, image analysis.

In our lab we study cell biology from a quantitative imaging standpoint. We use automated microscopy and image analysis to derive quantitative information from single cells, covering multiple cellular features and functional readouts. We employ a methodology to display these data simultaneously in cytometry-like scatter plots, which we have named QIBC (Quantitative Image Based Cytometry) [1,2] (Figure 1). This type of data representation serves as a visualization and discovery tool with great analytical capabilities. We use this methods on a daily basis, and it can also be applied to perform image-based screens with complex quantitative phenotypic readouts. We would like to share with the community how we exploit this technique in our particular field of study (DNA replication and DNA damage), hoping to rise interest and build future connections with other researchers.

Figure 1: Pipeline scheme and visual example of QIBC [1] Ochs, F. et al. 53BP1 fosters fidelity of homology-directed DNA repair. Nat Struct Mol Biol 23, 714â&#x20AC;&#x201C;721 (2016).

[2] Toledo, L. I. et al. ATR Prohibits Replication Catastrophe by Preventing Global Exhaustion of RPA. Cell 155, 1088â&#x20AC;&#x201C;1103 (2013)]

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-O4

Session: Imaging multicellular systems, Live imaging of single cells, Correlative Light and Electron Microscopy (CLEM) Nanoscopic Structure of Spider Silk Revealed by Super resolved Raman and Helium Ion Microscopy Irina Iachina1, Jacek Fiutowski2, Serguei Chiriaev2, Horst-GĂźnter Rubahn2 and Jonathan Brewer1

1) Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, 5230 Odense, Denmark.

2) Mads Clausen Institute, NanoSYD, University of Southern Denmark, Alsion 2, 6400 Sønderborg, Denmark *E-mail: brewer@bmb.sdu.dk Keywords: Spider silk, He Ion microscopy, CARS, Raman, super resolution.

Spider silk has multiple properties that are of industrial interest. For exsample it has a tensile strength comparable to that of alloy steel while being six times lighter and the silk is made of proteins at room temperature making it an environmentally friendly super material. Using multiple types of microscopy, we characterize Major Ampullate silk (MAS) and Minor Ampullate silk (MiS) spider silk fibers from the orb web weaving spider Nephila Madagascariensis to determine the, nano- and microscopic structures within the silk and couple these to the macroscopic properties such as tensile strength and elasticity.

Using Coherent Anti-Stokes Raman Scattering (CARS) and fluorescence microscopy the lipids and proteins of the fiber were analyzed and visualized revealing the overall structure of the fiber. To image the nanoscopic structures within the silk, He Ion and super-resolved Raman Microscopy was applied. By surface sputtering it was possible to etch away the outer most layers in order to visualize the inner protein arrangements with no special sample preparation. Using super resolved Raman microscopy the nanoscopic fibrils within the fibers could be imaged.

The combined He Ion and super resolution Raman images enable visualization of the nanoscopic stucure of the protein core of the fibers showing that it consists of fibrils arranged parallel to each other along the long axis of the fiber.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P1

Session: Imaging multicellular systems Fluorescence Imaging of Birch Mitosis and Meiosis Kesara Anamthawat-Jónsson*1 1

Institute of Life and Environmental Sciences, University of Iceland, 101 Reykjavík, Iceland. *E-mail: kesara@hi.is Keywords: Betula, chromosomes, hybridization, introgression, light microscopy.

The aim of this presentation is to demonstrate the importance of light and fluorescence microscopy in the study of plant genecology and population-based genetics [1].

Birch woodland is an integral component of the Arctic tundra. Tundra vegetation is mostly herbaceous, consisting of a mixture of mosses and lichens, grasses and forbs, along with dwarf shrubs and trees. Birch (Betula L.) is generally the most dominant woodland plant. Two species of Betula co-exist in Iceland: diploid dwarf birch (B. nana) and tetraploid downy birch (B. pubescens). They hybridize in their natural habitats, giving rise to triploid hybrids.

Analysing mitotic chromosomes of birch tree species has made it possible to detect interspecific hybridisation in natural woodlands, the process that drives gene flow across species boundaries. Hundreds of samples were collected from woodlands all around Iceland and the results of chromosome counting show that triploid hybrids are relatively common [2].

In addition to fluorescent staining of chromosomes from apical meristems, the technique of fluorescence in situ hybridisation (FISH) was used to map the major 45S ribosomal genes on birch chromosomes, further confirming the identity of hybrid and species (Fig. 1A).

Figure 1: A) Tetraploid Betula pubescens has six 45S ribosomal loci, including four actively transcribing ribosomal RNA into the nucleolus. B) A pollen mother cell of B. pubescens showing normal chromosome behaviour at diakinesis with 28 bivalents. Scale bar = 10 µm.

Capturing meiotic chromosomes in division, which occurs one week once a year, has confirmed the role of hybrids in mediating the gene flow. To demonstrate that triploid birch hybrids are able to produce fertile gametes, I investigated chromosome behaviour at meiosis in male catkins (Fig. 1B). The results show that the hybrids produce viable pollen and seeds.

Understanding the ecology and genetics of woodland birch can therefore help manage the ongoing conservation and regeneration of natural vegetation effectively.

[1] K. Anamthawat-Jónsson, Imaging & Microscopy 20(1), 18-20 (2018).

[2] K. Anamthawat-Jónsson, InTech – Open Access Publisher, pp. 117-144 (2012).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P2

Session: Imaging multicellular systems Substrates For Identification Of The Bone Marrow Mesenchymal Stem Cells By Surface-Enhanced Raman Spectroscopy Adrianna Milewska*1,2, Ólafur Eysteinn Sigurjónsson2,3,4, Milos Miljkovic5, Igor Sokolov5, and Kristján Leósson1,3.

1

2

Innovation Center Iceland, Árleynir 2-8, 122 Reykjavík.

The Blood Bank, Landspitali University Hospital, Snorrabraut 60, 105 Reykjavík.

3 University of Icealnd, Sæmundargötu 2, 101 Reykjavík.

4 Reykjavik University, Menntavegur 1, 101 Reykjavík.

5 Tufts University, 200 Boston Ave., Medford MA 02155, USA. *E-mail: adrianna@nmi.is

Keywords: surface-enhanced Raman scattering, mesenchymal stem cells, cell analysis.

The identification of bone marrow-derived mesenchymal stem cells (BM-MSCs), especially in non-destructive and non-invasive manner, has challenged researchers for decades.

Numerous methods have been utilized to characterize the MSCs, including chemical staining assays, quantitative polymerase chain reaction (qPCR) and immunofluorescence staining.

These methods are effective and relevant for the intensive analysis of the cells, but their assessment process is destructive and invasive. Therefore, a critical need has arisen to develop a new analytical method that enables sensitive non-destructive characterization of such stem cells without using any destructive steps. Surface-enhanced Raman spectroscopy (SERS) provides a potential platform to meet this need. SERS substrates have the ability to enhance Raman scattering by several orders of magnitude (typical enhancement factors of 106-108) when a molecular structure of the cell is located in close vicinity of nanostructured noble metal surfaces such as Au or Ag. Herein, we present an investigation of BM-MSCs grown on novel cell-compatible SERS substrates prepared by using a repeated gold deposition and thermal annealing method. MSCs were cultured on the substrates for 7 days and demonstrated increasing proliferation and low cytotoxicity level. Fluorescence staining confirmed cell attachment to the surface and SERS measurements demonstrated Raman peaks that can be associated with actin filament-building proteins. Preliminary results suggest that the SERS technique can provide a sensitive and nondestructive method for studying mesenchymal stromal cells.

Figure 1: (a) SEM image of SERS substrate surface. (b) Fluorescence microscopy of focal adhesion, actin cytoskeleton and nuclei in MSCs on a SERS substrate. (c) Optical microscope image overlaid with the Raman spectroscopy image (462.4 cm-1 band). (d) SERS Raman spectra recorded at several points on the image [1] X. Sun, H. Li, Nanotechnology, 2013, 24(35):355706.

[2] X. Cao, Y. Shan, L. Tan, X. Yu, M. Bao, W. Li, H. Shi, J. Mater. Chem. B, 2017, 5, 59835995.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P3

Session: Imaging multicellular systems, Live imaging of single cells, Correlative Light and Electron Microscopy (CLEM) Life Cycle of Bdellovibrio bacteriovorus imaged by Helium Ion Microscopy Nedal Said*1, Matthias Schmidt1, Markus Krüger2, Antonis Chatzinotas2 Helmholtz Centre for Environmental Research – UFZ, 1 - Department of Isotope Biogeochemistry, ProVIS – Centre for Chemical Microscopy 2- Department of Environmental Microbiology.

Email: Nedal.Said@ufz.de Abstract Bdellovibrio bacteriovorus HD100 is an obligate predatory bacterium which requires live Gram-negative bacteria as prey for growth and replication The growth cycle usually includes an extracellular attack phase in which the predatory bacterium hunts for its prey and invades the prey cell, followed by an intracellular replication phase, during which the invaded prey cell is transformed into a bdelloplast. Finally, the destruction of the prey cell leads to the release of two to seven new Bdellovibrio cells. In this study we employed high-resolution helium-ion microscopy (HIM) to image Bdellovibrio in attack phase, during invasion and penetration of the prey and after the bdelloplasts, i.e. the invaded cells containing the offspring, had formed. The investigations were carried out with two prey species, Escherichia coli and Pseudomonas putida. For that a preparation protocol involving chemical fixation, filtration, post-fixation and dehydration that preserves the morphology of both, Bdellovibrio and prey, were developed.

The experiments provided evidence for multiple invasions per prey cell. The novel preparation of the bacteria as well as the high-resolution of the HIM allowed to observe the morphological changes that take place in the prey cells after invasion. These studies provide a means to image bacterial predation at high-resolution which in future can be transferred to environmental predator-prey systems.

Helium ion micrograph of a Bdellovibrio invading an E.coli.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P4

Session: Imaging multicellular systems Efficiency of Antimicrobial Modified Hyperbranched Polyethyleneimine Polymers Analogous With Different Microstructures Mériem Er-Rafik*1,2, YingChun He2, N. Pasquier2, H. Keul2 and M. Moeller2 1

Institut Charles Sadron, Université de Strasbourg-CNRS UPR 22, 67034 Strasbourg, France 2 DWI Interactive Material Research, RWTH Aachen University, Germany *E-mail: meriem.er-rafik@ics-cnrs.unistra.fr

Keywords: Biomimetic antimicrobial polymer, permeability, microstructure Antimicrobial synthetic polymers are in demand for industrial applications like disinfection and antimicrobial coatings. Ultimately antimicrobial polymers may be useful antibiotics even in pharmaceutical applications, depending on the range of targeted organisms, and the absence of hemolytic activity (destruction of mammalian red blood cells).

The physicochemical principles by which antimicrobial peptides adhere to the cell envelope and finally destroy its integrity might be implemented in the design of synthetic macromolecules. Although, the structure property relationships yielding an antimicrobial polymer, three elements are obviously beyond any doubt: (i) the polymer must be available in the aqueous medium, in which microbes proliferate; (ii) the polymer must contain hydrophobic elements to be attached on or integrated in the cell membrane, and (iii) cationic charges promote attack of the cell envelope and selectivity for microbes. Here water soluble hyperbranched polymers can be expected to provide a readily available alternative to design amphipathic structures, mimicking some of the features discussed for antimicrobial peptides.

Cationic amphiphilic polymers were prepared from poly(ethylene imine) (PEI) and functional ethylene carbonates bearing cationic, hydrophobic or amphiphilic groups [1]. The polymers are designed to exhibit antimicrobial properties. In a one-step addition different functional ethylene carbonates were added to react with the primary amine groups of PEI. The water soluble polymers were studied regarding their ability to form soluble aggregates, their antimicrobial activity in relation to the ratio of alkyl/cationic groups, length of the alkyl chains, molecular weight of the PEI against E. coli, their permeability potential and the effect on the ultrastructure of E. coli.

Figure 1: Centered figure in line with text, with explaining caption.

[1] N. Pasquier et al., Macromolecular Bioscience, 8, 903–915 (2008).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P5

Session: Live imaging of single cells High-throughput screening for mitotic functions of TopBP1 Michael Lisby*1,2, Jonas Bagge1, Nanna KorsbĂŚk Smedengaard1 and Vibe H. Oestergaard1 1

Department of Biology, University of Copenhagen, Ole Maaloees Vej 5, 2200 Copenhagen N, Denmark.

2 Center for Chromosome stability, Department of Cellular and Molecular Medicine, University of Copenhagen, Blegdamsvej 3b, 2200 Copenhagen N, Denmark. *E-mail: mlisby@bio.ku.dk Keywords: mitosis, chromosome segregation, DNA repair.

Genome integrity is critically dependent on timely DNA replication and accurate chromosome segregation prior to cell division. TopBP1 is a multi-functional scaffold protein with roles in DNA replication, DNA damage checkpoint signalling and recombinational DNA repair, where it facilitates the formation of transient protein complexes. Recently, we have shown that TopBP1 also plays a crucial role in mitotic segregation of chromosomes. Specifically, we found that TopBP1 is required for recruitment of the structure-selective nuclease SLX4 to chromatin, which may promote resolution of DNA repair intermediates, and TopBP1 localizes to ultrafine DNA anaphase bridges (UFBs) that form, when the cell attempts to segregate intertwined sister chromatids. By using precise temporal depletion of TopBP1 just prior to mitotic entry, we could demonstrate that mitotic cells rely on TopBP1 to prevent formation of 53BP1 nuclear bodies in the next cell cycle, showing that TopBP1 acts to reduce transmission of DNA damage to G1 daughter cells. Consistently, depletion of TopBP1 also lead to an increased frequency of mitotic failure (binucleated cells), defective mitotic DNA synthesis (MiDAS), and accumulation of chromosome bridges in mitosis. Based on these results, we propose that TopBP1 maintains genome integrity in mitosis by controlling mitotic DNA repair and chromosome segregation. To identify the underlying molecular mechanisms by which TopBP1 mediates faithful mitotic inheritance of chromosomes, we are currently screening a library of FDA-approved therapeutics for compounds that activate or inhibit the TopBP1 pathway.

Figure 1: A DNA ultra-fine anaphase bridge connecting segregating chromosomes (in red) is bound by the PICH translocase (in blue) and the TopBP1 scaffold protein (in yellow) (Germann et al. 2014).

References Germann SM, Schramke V, Pedersen RT, Gallina I, Eckert-Boulet N, Oestergaard VH, Lisby M. 2014. TopBP1/Dpb11 binds DNA anaphase bridges to prevent genome instability.

J Cell Biol 204: 45-59.

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P6

Session: Imaging multicellular systems, Live imaging of single cells, Correlative Light and Electron Microscopy (CLEM) Structome analysis and three-dimensional reconstitution of Mycobacterium smegmatis cells Hiroyuki Yamada*1, Masashi Yamaguchi2, Kinuyo Chikamatsu1, Akio Aono1, Nagatoshi Fujiwara3, Yuriko Igarashi1, Akiko Takaki1, and Satoshi Mitarai1.

1 2

Research Institute of Tuberculosis, JATA., Kiyose, Tokyo, 204-8533, Japan,

Medical Mycology Research Center, Chiba University, Chiba, 260-8673, Japan 3

Tezukayama University, Nara 631-8585, Japan

*E-mail: hyamada@jata.or.jp Keywords: Mycobacteria, quick freezing, freeze-substitution, serial ultrathin sectioning, TEM We have already reported the results of structome analysis, “quantitative and three dimensional structural information of a whole cell at the electron microscopic level,” of Exophiala dermatitidis, Saccharomyces cerevisiae, Mycobacterium tuberculosis (MTB), Escherichia coli and Myojin spiral bacteria. In this study, structome analysis of M. smegmatis (MSG), nonpathogenic mycobacterium, have been performed to compare especially with highly pathogenic MTB as well as other species above.

Materials and methods: MSG was cultured with Middlebrook 7H9 broth containing 0.05% Tween 80 and supplemented with OADC enrichment for 1 week. After centrifuged at 10,000 x g for 1 min, the pellet was subjected to quick freezing with Vitrobot Mark IV (FEI) and to freeze-substitution with 2% osmium tetroxide-acetone solution at -80 °C for several days.

Then, the sample were rinsed with absolute acetone and embedded with Spurr’s resin. Totally 233 serial ultrathin sections of 50 nm thickness were cut and seven cells in the sections were examined with JEOL JEM-1230. Images were analyzed with ImageJ/Fiji and cell diameter (outer membrane; OM, and plasma membrane; PM), cell volume (OM/PM), cytoplasmic ribosome number and ribosome density are measured and calculated.

Results and discussion: Average ± SD of diameter (OM/PM), volume (OM/PM), total ribosome number and ribosome density per 0.1 fl cytoplasm were 0.587 ± 0.055 μm / 0.550 ± 0.051 μm, 1.13 ± 0.46 fl (μm3) / 0.99 ± 0.39 fl, 8,670 ± 2,660, and 910 ± 143, respectively.

Compared with MTB, MSG cells are significantly larger (p < 0.002), contains significantly more cytoplasmic ribosomes (p < 0.0002) and have significantly higher cytoplasmic ribosome density (p < 0.05). These results support that MSG cells can grow faster than MTB cells.

Figure 1: Ultrathin section image of quick-freeze and freeze-substituted MSG cell.

[1] S.K. Biswas, et al., J. Electron Microscopy (Tokyo), 52, 133–143 (2003).

[2] M. Yamaguchi, et al., J. Electron Microscopy (Tokyo), 52, 133–143 (2011).

[3] H. Yamada, et al., PLoSOne, 10, e0117109 (2015).

[4] M. Yamaguchi, et al., Microscopy (Oxf), 65, 363–369 (2016).

[5] H. Yamada, et al., Microscopy (Oxf), 66, 283-294 (2017).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P7

Session: Imaging multicellular systems, Live imaging of single cells, Correlative Light and Electron Microscopy (CLEM) CLEM and SEM in inflammatory responses M. Baumgartenยง, R. Bhongirยง, S. Hari*, H. Herwaldยง ยงLund University, Division of Infection Medicine, IQ Biotechnology Platform *Delmic B.V., The Netherlands

* E-mail: maria.baumgarten@med.lu.se Keywords: CLEM, SEM, desktop, correlative microscopy, inflammation.

Microscopy is an important tool to detect inflammatory reactions at cellular level. Here we report that correlative light and electron microscopy (CLEM) can be used to study morphological changes of immune cells under ex vivo conditions. Our data also show that CLEM can be employed to investigate our animal models of infection. Together our data reveal that CLEM can be used to for studying host responses under in vitro, ex vivo, and in vivo conditions.

The Delphi is an all-in-one solution for correlative light and electron microscopy (CLEM).

It is an integrated desktop scanning electron microscope (SEM) including an inverted widefield fluorescence microscope. This integration enables scientists to perform correlative microscopy without the challenges typically associated with CLEM. Here we examine the modulation of inflammatory responses to bacterial infections.

Neutrophils play an important role during infection and inflammation. Once activated neutrophils change their morphology and release their content including antimicrobial substances, reactive oxygen species, and vasodilating substances. Notably, our previous work has shown that microscopes are an important tool to monitor bacteria-induced tissue damage and counteracting host responses.

This study aims to identify and characterize molecular mechanisms that lead to such complications.

CLEM will provide better results as the fluorescence-targeted proteins will show the exact locations of the structures of interest..

Figure 1: The CLEM images can be used to identify the neutrophil status

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P8

Session: Imaging multicellular systems Ultrastructural Dynamics of Podospora anserina Mycelium under Conditions of Long-Term Evolutionary Experiment Olga A. Kudryavtseva*1 and Igor S. Mazheika1.

1

Department of Mycology and Phycology, Faculty of Biology, Lomonosov Moscow State University, Moscow, Russia. *E-mail: kudryavtseva@mail.bio.msu.ru Keywords: experimental evolution, fungal model, electron microscopy, large doublemembrane vesicles.

The present study examines ultrastructural changes observed in filamentous fungus Podospora anserina in the course of long-term laboratory evolution. Our scientific group succeeded in finding experimental conditions that triggers irreversible changes in P. anserina phenotype. Acceptable conditions are continuous mycelia cultivation in liquid nutrient medium with the use of orbital shaker [1]. We furthermore demonstrated positive selection of de novo mutations in our model system [2]. Despite the fact that most of acquired mutations belong to different genomic sites, phenotypic changes cover the same morphological aspects in independent P. anserina lines. Eight lines derived from two initial P. anserina strains of wild type were removed from agar containing to liquid medium in the year of 2012. Since then continuous growth of all lines is maintained by simultaneous serial passages. The evolutionary experiment is still in progress.

Ultrastructural study let us conclude that during the short period right after the experiment initiation P. anserina subjected to the stress, but after that adapted to new conditions of growth. Fungal cells in the course of short adaptation phase had thick double layered or multilayered cell walls and additionally were protected with fibrillous covers. Cytoplasm demonstrated high electron density making it quite difficult to trace intracellular components.

Mitochondria showed rounded profiles, they were small and numerous probably because of fragmentation. Nuclei were comparatively small. Mycelia adaptation was characterized by dramatic increase of biomass yield. At the ultrastructural level adapted lines had some distinctive features, the most important of which are: thin cell walls (mainly single-layered) having no additional protection, big nuclei which diameter frequently was almost equal to hyphal width, elongated mitochondrial profiles with parallel cristae. Electron density of cytoplasm decreased making perfectly visible all cell structures.

Two and a half years after experiment start the new distinctiveness of P. anserina intracellular life was noticed. Cellular vacuoles became to capture, absorb and degrade huge parts of cytoplasm that is not usual for P. anserina wild type strains. It is possible that this is the way for the fungus to realize recycling of intracellular materials faster.

The most surprising characteristic of P. anserina under conditions of growth in liquid medium was formation of special double-membrane vesicles. Their length ranges between 130 and 760 nm, width varies between 100 and 500 nm (the average values are 336Âą20 nm and 222Âą12 nm respectively). The experiment continuation led to enrichment of P. anserina cells by the vesicles of such kind. After five years of growth in liquid large double-membrane vesicles became common for experimental lines as a result of which we could follow their way of formation. Plasma membrane invaginates very deeply into cytoplasm, curves and forms hemisphere with electron-translucent content (Fig. 1). After closure the vesicles migrate into different parts of cell cytoplasm. In fungi double-membrane vesicles originated from the cell membrane were occasionally noticed by classical [3] and modern [4] authors, but their

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P9

Session: Imaging multicellular systems functional purpose is debatable and essentially unknown. It may be more delicate and sophisticated mechanism of recycling. Mitochondria often could be found near forming double-membrane vesicle indicating that this process may need some energy.

P. anserina mycelial isolates being transferred from adapted experimental cultures to solid medium (standard conditions) demonstrates all modifications acquired by the appropriate submerged culture. On the level of ultrastructure isolates are indistinguishable from submerged mycelia.

The reported study was funded by RFBR, research project № 18-04-01349 a.

Figure 1: Formation of large double-membrane vesicle in P. anserina experimental line (2,5 years under evolutionary experiment), fixation with glutaraldehyde and osmium tetroxide. CW – cell wall, Pm – plasma membrane, Cy – cytoplasm.

[1] O.A. Kudryavtseva et al., Microbiology 80:6, 784–796 (2011).

[2] O. Kudryavtseva et al., Journal of Microbial & Biochemical Technology: Applied & Food Microbiology 9:6(Suppl), 48 (2017).

[3] R.C. Aylmore and N.K. Todd, The Ecology and Physiology of the Fungal Mycelium.

Edited by D.H. Jennings and A.D.M. Rayner, 103–126 (1984).

[4] R.Jr. Taylor et al., Autophagy 8:9, 1300–1311 (2012).

SCANDEM 2018, Technical University of Denmark, Kgs. Lyngby, Denmark H-P9

Author Index A Adamsen, K. C. C-02 Ahmed, F. K-3 Alekseeva, S. C-01, A-P14 Alm, K. H-01, G-P7 Anamthawat-Jónsson, K. H-P2 Andersen, I. M. G-P1 Andersson, M. F-I1 Andresen, T. L. H-P8, F-02 Antosiewicz, T. J. C-01 Aono, A. H-P7 Arlinghaus, H. B-P2 Asokan, V. A-P9 Ayata, M. G-P2 B Baier, S. F-01 Bagge, J. H-P6 Balantekin, M. F-P3 Baumgarten, M. H-02 Bech, M. F-I1 Beleggia, M. E-04, E-P1,

D-03

Belevich, I. G-01 Bencan, A. A-I1 Bewick, A. E-P2 Bhongir, R. H-O2 Bigdeli, S. A-P9 Blazit, J. D. E-I1 Bon, M. B-P4 Boni, A. H-I2 Bornhoefft, M. C-O3 Brar, H. S. F-P3 Breivik, T. A-P15 Brewer, J. H-P1 Brezesinski, T. B-P14 Brostrøm, A. G-P4, D-O4 Burrows, A. E-O3, C-O1, A-P14 Busam, J. A-P3 Busse, D. A-P5 Bystron, A. M. G-O5 Bøjesen, E. D. G-O2 C Cacho-Nerin, F. F-P2 Campos A. E-I1 Chatzinotas, A. H-P4 Chen, F.-R. B-O3

Chen, M. A-P18 Chikamatsu, K. H-P7 Chiriaev, S. H-P1 Chorkendorff, I. P-1, F-O3, A-P13, B-P1 Christensen, J. M. A-O1 Colding-Jørgensen, S. A-P12 Collinson, L. K-1 Colvin, J. F-O4 Conradsen, K. G-P5 Creemer, J. F. B-P1 D Dahl, A. B. F-I1, G-O6, G-P5 Damjanovic, D. A-I1 Damsgaard, C. D. F-O1, A-O1, B-P1 Darakchieva, V. C-O6 de Medeiros, G. H-I2 Deepak, F. L. A-O2 Del Cerro, P. E-O1 den Dekker, A. J. G-O4 Delimitis, A. G-O3 Dianoux, R. B-P2 Dick, K. A. F-O4, B-O2 Dicks, K. E-P2 Dimitriadis, D. A-P8 Diplas, S. A-P6 Dluhus, J. C-O3 Dona, P. B-P1 Drazic, G. A-I1 Duetting, A. B-P2 Dyrby, T. B. F-I1 E Ehrlich, N. H-P8 Ek, M. B-O3 Eklund, J. A-P9 Elkjær, C. F. B-P1, B-P9 Elm, M. T. B-P14 Emerson, M. J. G-P5 Engelhardt, P. A-P16 Elsukova, A. E-O4, E-P1, D-O3 Er-Rafik, M. H-O3, H-P5 Erni, R. B-O4, B-P4 Escrig, S. A-P8 Etheridge, J. G-O2

F Fanta, A. B. d. S. E-O3, C-O1, C-O5, A-P1, A-P14 Fatermans, J. G-O4 Ferji, K. H-O3 Fester, J. A-P7 Fimland, B.-O. G-P1 Fiutowski, J. H-P1 Flyvbjerg, H. H-P8 Frondelius, T. A-P4 Fujiwara, N. H-P7, G-P2 G Gamaletsos, P. N. A-P8 Gaulandris, F. B-P6 Gavrilowicz, G. G-P6 Geppert, J. A-P5 Ghadimi, R. B-P7 Gjønnes, J. G-O3 Gloter A. E-I1 Godelitsas, A. A-P8 Gouillart, E. K-5 Gram, L. P-2 Grumsen, F. A-P10 Grunwaldt, J.-D. F-O1 H Halim, J. C-O6 Hammer, B. C-O2 Han, A. E-O4, E-P1, C-O5, D-O3 Hansen, L. P. B-O3 Hansen, P. C. G-O6 Hansen, T. W. C-O6, A-O1, B-P8 Hansen, V. G-O3 Hari, S. H-O2 Harlin, P. C-O4 He, YC. E-P1 Heinig, M. F. A-P1 Helveg, S. B-O3, B-P1, B-P9 Hempel, C. F-O2 Hendriksen, B. B-P1 Henninen, T. B-O4, B-P4 Herwald, H. H-O2 Hintikka, J. A-P4 Hjelen, J. A-P15 Hofer, C. B-P10 Honkanen, M. A-P2, A-P4 Horch, S. F-O3 Huang, X. H-O1 Hussein, A. B-P10

Huuhtanen, M. A-P2 Hyllested, J. Æ. B-P11 I Iachina, I. H-P1 Iandolo, B. C-O1, A-P14 Idrobo,. J.-C. E-O2 Igarashi, Y. H-P7 Ikkala, O. A-P16 J Jacobsson, D. B-O2 Janicke, B. H-O1 Jansen, H. C-O5, A-P1 Jensen, A. D. A-O1 Jensen, F. B-P11 Jensen, I. J. T. A-P6 Jensen, L. H. S. A-P8 Jeppesen, N. G-P6 Jespersen, K. M. G-P5 Jespersen, S. P. B-P1 Jiang. H. A-P2, A-P11 Jinschek, J. R. B-O3, B-P1 Johansson, J. B-O2 Jokitalo, E. G-01 Jones, L. E-02 Jonsson, T. A-P9 Jooss, C. A-P5 Juoksukangas, J. A-P4 K Kadkhodazadeh, S. A-P1 Kallinen, K. A-P2 Kamlund, S. H-01 Kapishnikov, S. F-02 Karlsen, M. A-P15 Kasama, T. A-P8, A-P14, B-P11 Kauppinen, E. I. A-P11, B-P10 Kayser, S. B-P2 Keiski, R. L. A-P2 Keller, D. B-O4 Keul, H. H-P5 Kibsgaard, J. A-P13 Kisielowski, C. B-O3 Kjeldstad, T. A-P6 Kjer, H. M. F-I1 Kleppen, J. A-P15 Klímek, P. C-O3 Kling, K. I. G-O5, G-P4 Knutsson, J. V. F-O4

Kociak, M. E-I1 Kollmer, F. B-P2 Kolsbjerg, E. L. C-O2 Kon, Y. A-P8 Kotakoski, J. D-O1, B-P10 Koo, J. G-O6 Kooyman, P. B-P1 Koust, S. C-O2 Krüger, M. H-P4 Kudryavtseva O.A., H-P9 Kuhn, L. T. A-P12, B-P6 Kuwata, H. G-P2 Kylberg, G. G-P3 Kärkkäinen, M. A-P2 L Langhammer, C. C-O1, A-P14 Lauritsen, J. V. K-2, F-P1, C-O2, A-P7, B-P13 Lautrup, L. C-O4 Le Meins, J.-F. H-O3 Lehtovaara, A. A-P4 Leósson, K. H-P3 Li, J. A-O2 Li, X. E-I1 Li, Y. B-P13 Liberali, P. H-I2 Lisby, M. H-P6 Liu, A. C. Y. G-O2 Liu, P. B-P8 Liu, Y. F-O4 Liu, Y. A-P18 Liu, Y.-P. F-O4 Lobato, I. G-O4 Lourenco-Martins E-I1 Ludacka, U. D-O1 Lukonin, I. H-I2 Løvvik, O. M. A-P6 M Maagaard, T. F-O3 Madsen, J. B-P8 Maeda, S. G-P2 Malereanu, R. A-P1 Maletta, M. E-P3 Maliakkal, C. B. B-O2 Marques, C. M. H-O3 Mayr, U. H-I2 McKibbin, S. R. F-O4 Meibom, A. A-P8

Mele, L. B-P1 Meuret, S. E-I1 Meyer, J. C. D-O1, B-P10 Michel, K. B-P14 Michler, J. A-P19 Mikkelsen, A. F-O4 Mikkelsen, L. P. G-P5 Milewska, A. H-P3 Miljkovic, M. H-P3 Mirsaidov, U. M. D-I1 Mirzayev, R. B-P10 Mitarai, S. H-P7 Moeller, M. H-P5 Moellers, R. B-P2 Mohanty, G. A-P19 Monazam, M. R. A. D-O1, B-P10 Mortensen, K. I. H-P8, D-O4 Muggerud, A.-M. F. A-P3 Mustonen, K. B-P10 Muto, S. B-P6 Müller-Caspary, K. G-O4 Mäntylä, A. A-P4 Mølhave, K. G-P4, D-O4, B-P5, B-P6 Møller, P. A-P10 N Naka, T. G-P2 Negi, D. S. E-O2 Niessen, F. E-O3 Nielsen, J. O. A-P10 Nielsen, M. R. A-O1, B-P8 Niehuis, E. B-P2 Nilsen, J. S. G-P1 Nonappa A-P16 Nordby, T. B-P14 Nugroho, F. A. A. C-O1 Nurmi, V. A-P4 Näslund, L.-Å. C-O6 O Oestergaard, V. H. H-P6 Ormstrup, J. F-P4 Oredssson, S. H-O1 Oster, M. B-P7 P Pabitra, D. E-I1 Pacureanu, A. F-I1 Palisaitis, J. C-O6 Pantleon, K. A-P10

Papasaikas, P. H-I2 Parker, J. E. F-P2 Pasquier, N. H-P5 Passerone, D. B-P4 Paulsen, R. R. G-P6 Persdotter, A. A-P9 Persson, A. R. B-O2 Persson, I. C-O6 Persson, P. O. Å. C-O6 Petersen, T. C. G-O2 Petit, L. E-O1 Phifer, D. E-P4 Q Qiu, M. E-P1 Qvortrup, K. F-O2 Quinn, P. D. F-P2 R Rading, D. B-P2 Ramasse, Q. K-4 Raza, S. D-O2 Ren, D. G-P1 Rentenberger, C. D-O1 Risch, M. A-P5 Roddatis, V. A-P5 Rodrigues-Fernandez, J. A-P7 Rojac, T. A-I1 Roma, G. H-I2 Rosén, J. C-O6 Rubahn, H.-G. H-P1 Rusz, J. E-O2 Ryner, M. G-P3 S Said, N. H-P4 Salminen, T. E-O1 Sandre, O. H-O3 Sanna, S. B-P6 Scheidemann, A. B-P2 Schiøtz, J. B-P8 Schmidt, M. H-P4 Schmidt, S. A-P12 Schmutz, M. H-O3 Schumann, M. A-O1 Secher, N. M. A-P13 Sehested, J. B-P9 Sellergren, B. G-P7

Semenova, E. A-P14 Serra, D. H-I2 Shihavuddin, A. G-P6 Shinde, S. G-P7 Sigurjónsson, Ó. E. H-P3 Simonsen, S. B. A-P12, B-P6 Sintorn, I.-M. G-P3 Sláma, M. C-O3 Smedengaard, N. K. H-P6 Snellman, A. A-P17 Sokolov, I. H-P3 Stadler, M. H-I2 Stange, M. A-P6 Stéphan, O. E-I1 Sternbæk, L. G-P7 Strnad, P H-I2 Sukham, J. A-P1 Sun, A. A-P7 Sun, C. B-P12 Sun, H. D-O4, B-P5 Sun, X. A-P18 Suomela, J. A-P17 Susi, T. D-O1, B-P10 Sølling, T. I. G-O5 Sørensen, B. E. A-P15 T Tian, Y. A-P11 Takagi, T. A-P8 Takaki, A. H-P7 Tan, T. A-P7 Tang, M. B-P3 Tassone, C. H-P8 Tencé, M. E-I1 Tesarová, H. C-O3 Thøgersen, A. A-P6, B-P14 Tiddi, W. E-O4, E-P1 Tidemand-Lichtenberg, S. D-O4 Tietz, D. B-P7 Tietz, H. B-P7 Timm, R. F-O4 Tizei, L. H. G. E-I1 Todeschini, M. C-O5, A-P1 Toledo, L. H-O4 Tornberg, M. B-O2 Trojan, A. F-O4

V Vajanto, K. A-P17 Van Aert, S. G-O4 van den Berg B-P9 van Dyck, D. B-O3 van Helvoort, A. T. J. G-P1, A-P3 Vendelbo, S. B-P1 Viazmitinov, D. A-P14 Vippola, M. A-P2, A-P4 Vojvodic, A. A-P7 Volkmann, K. H-I2 W Wagner J. B. C-O1, C-O5, C-O6, A-O1, A-P1, A-P10, B-P6, B-P8, B-P11 Waldt, A. H-12 Wallenberg, R. B-O2 Walter, T. H-I1 Wang, Y. G-P5 Wang, Y. B-O1, B-P3, B-P12 Webb, J. L. F-O4 Weman, H. G-P1 Wendt, S. F-P1, C-O2 Wenner, S. A-P3 Wenzell, B. A-P8 Westermann, I. A-P15 Wingren, A. G. G-P7 Wisnet, A. B-P7 Withers, P. J. G-P5 X Xu, T. F-P1

Y Yamada, H. H-P7, G-P2 Yamaguchi, M. H-P7 Yesibolati, M. N. D-O4, B-P5, B-P11 Yngman, S. F-O4 Yu, Y. A-P15 Yuan, W. B-P12 Z Zhang, L. A-P7 Zhang, M. B-P5 Zhang, W. A-P12 Zhang, Z. B-P12 Zhdanov, V. C-O1 Zhao, D. E-O4, E-P1 Zhu, B. B-P3 Zhu, L. G-P3 Zobelli A. E-I1 Zunic, T. B. A-P8 Ă&#x2026; Ă&#x2026;nes, H. W. G-P1

We thank all our corporate partners for fueling Scandem 2018

303 A - 27

303 A - 28

303 A - 24

306 - 3

306 - 6

303 A - 30

306 - 15

306 - 5

303 A - 23

303 A - 26

303 A - 34

306 - 16

306 - 18

306 - 13

306 - 21

306 - 20

306 - 22

306 - 6

303 A - 25

306 - 4

303 A - 31

306 - 19

306 - 2

306 - 4

306 - 14

303 A - 33