Tecan Journal Edition 01/2013 by Tecan Group Ltd.

Tecan Journal Life Sciences and Partnering Business

Edition 1/2013

Baked to perfection pages 12-13

More than skin deep page 19

Rewriting the proteomics handbook pages 24-25

Automated screening for recessive disorders elevates sensitivity to new heights pages 30-31

CEO WELCOME TECAN JOURNAL 1/2013

Welcome to the first Tecan Journal of 2013

Welcome

Let me start by saying how delighted I am to be joining Tecan at this exciting time. Having been a Tecan customer for many years, I understand that innovation, quality and trust are cornerstones of the Company’s success, and look forward to working with colleagues and customers to ensure that Tecan remains the partner of choice for cutting-edge laboratory automation solutions. The SLAS2013 Exhibition in Orlando, Florida, once again provides us with an opportunity to demonstrate Tecan’s knowledge and broad product portfolio, showcasing an array of products which allow end-toend automation and optimization of our customers’ workflows. Highlights at this show include the latest TouchTools™ 3.0 user interface, which will aid the development and consistent implementation of laboratory automation, and the Tecan Academy, an exciting new training portal that will soon offer on-demand, web-based courses for operators of our liquid handling and

detection systems. The Infinite® 200 PRO with Gas Control Module (GCM™) will also be on display, providing an ideal solution for customers interested in cell-based assays, as well as our broad portfolio of Tecan Cavro® brand components for OEM customers. All of these solutions are designed to improve productivity and uptime by catering to the needs of our customers. As we see it, our job is to simplify yours! As well as details of these latest innovations, this issue of the Tecan Journal also includes a wide range of articles covering everything from speeding up drug discovery and the etiology of pancreatic cancers to the development of new ingredients for the baking industry and cosmetics. These diverse applications demonstrate how you, our customers, are using Tecan solutions to push the boundaries of science. I hope you enjoy the journal, and I look forward to a prosperous year for Tecan and our customers. David Martyr, CEO

CONTENTS TECAN JOURNAL 1/2013

page 6-7

Historic Boston plays host to the fifth Tecan Symposium 2

CEO Welcome

4 Talk to us @ SLAS2013

pages 10-11

5 AlphaScreen速 option launched for Infinite F200 PRO multimode reader 5 First Air LiHa modules shipped to customers 6 - 7 Historic Boston plays host to the fifth Tecan Symposium 8 - 9 Introducing David Martyr 10 - 11 A watchful eye on biosecurity 12 - 13 Baked to perfection 14 - 15 Speeding up drug discovery 16 - 17 Developing a proteomic fingerprint for pancreatic cancer 18 Made-to-measure robotics 19 More than skin deep 20 - 21 Too gracious a host

pages 22-23

22 - 23 Feeding the SRMAtlas project

pages 30-31

utomated screening A for recessive disorders elevates sensitivity to new heights

Contents

24 - 25 Rewriting the proteomics handbook 26 - 27 Undergraduate studies enter a whole new world 28 - 29 Investigating protein targets and cellular pathways in yeasts 30 - 31 Automated screening for recessive disorders elevates sensitivity to new heights 31 Leading the debate 32 Events

product NEWS TECAN JOURNAL 1/2013

Talk to us @ SLAS2013 Tecan innovation is everywhere, from state-of-the-art pharmaceutical development facilities to the smallest academic laboratories. As we see it, our job is to simplify yours, so visit Tecan on booth #701 at SLAS2013 to see how our comprehensive range of laboratory automation, detection and OEM liquid handling solutions could benefit your laboratory. TouchTools™ 3.0 – fully customizable touchscreen control The latest generation of Tecan’s TouchTools touchscreen user interface makes programming and operating Freedom EVO® workstations easier than ever before. TouchTools 3.0 introduces a fully customizable user interface, offering greater flexibility for instrument programmers and a structured, user-friendly environment for operators.

TouchTools 3.0 gives programmers the tools to create custom Maintenance and Run-Time workflows. New features make it easy to build tailored input screens, give set-up tips, and display step-by-step graphical instructions. These tools help to ensure smooth day-to-day operation with fewer operator errors. Creating assay-specific interfaces allows every aspect of a run to be described in terms familiar to laboratory scientists, including prompts, status updates, and user input screens. Embedding laboratory SOPs into the protocols will help improve compliance, enhance process security and provide traceability. TouchTools 3.0 has a new look and feel that echoes the latest consumer touchscreen devices. This intuitive interface makes experimental set-up quick and easy, and simplified workflows reduce the time required to train new operators.

Tecan Academy – on-demand, web-based training Going beyond traditional training courses, Tecan will soon be introducing an online training platform to provide instant access to user training and certification for various Tecan products. The Tecan Academy will offer users an easy way to learn new skills at a time and pace that suits their individual needs. Due to be launched in the US in Spring 2013, this straightforward web-based tool will enable participants to get up to speed faster, and ensures more consistent, error-free operation of Tecan hardware and software.

Infinite® 200 PRO – enhanced cell-based assays The patent pending GCM™ (Gas Control Module) for the Infinite 200 PRO is a breakthrough for cell-based experiments, allowing simultaneous control of atmospheric O2 and CO2 levels within the reader’s measurement chamber for the first time. Combined with the instrument’s continuous shaking and heating functions, this makes the Infinite 200 PRO the first hybrid incubator/reader on the market. Recent studies have demonstrated the system’s 3D assay capabilities using InSphero’s organotypic microtissue model, and artefacts resulting from evaporation can be minimized by using Thermo Scientific’s innovative Nunc™ Edge 96-well plate, offering a truly comprehensive solution for cell-based studies.

To find out more about these exciting new releases, visit Tecan on booth #701 at SLAS2013

product News TECAN JOURNAL 1/2013

AlphaScreen® option launched for Infinite F200 PRO multimode reader The newly developed AlphaScreen module for the popular Infinite F200 PRO multimode reader has been designed to provide customers with a cost-effective solution for AlphaScreen and AlphaLISA® assays. Offering excellent sensitivity for screening and research, the new option further extends the flexibility of the Infinite F200 PRO for low and medium throughput applications.

The bead-based AlphaScreen assay system

The AlphaScreen module combines the Infinite F200 PRO’s established fluorescence top optics with a bespoke filter set designed to use the instrument’s standard flashlamp. This set-up ensures optimal assay performance without the need for a specialized reader, complementing the

system’s absorbance, fluorescence intensity, luminescence, fluorescence polarization and time-resolved fluorescence resonance energy transfer measurement modes. To find out more on Tecan’s AlphaScreen solutions, visit www.tecan.com/alphascreen

A typical AlphaScreen signal curve using an Infinite F200 PRO with PerkinElmer’s P-Tyr-100 phosphotyrosine kinase detection kit

First Air LiHa modules shipped to customers Tecan is pleased to announce that the first Air LiHa air displacement pipetting arms have left Tecan’s production facility in Männedorf. Designed to offer an unrivalled level of flexibility for Freedom EVO liquid handling workstations, this innovative module is Tecan’s most compact single channel pipetting option, and offered a number of fresh challenges for the manufacturing team. Kaspar Städeli, Assembly Team Leader, commented: “The Air LiHa is a very compact unit. Like a good Swiss watch, many small parts need to work in perfect unison, and we have invested many hours in training and in discussion with our trusted suppliers to guarantee the highest quality standards. We have created a whole new generation of manufacturing tools and testing equipment for this project, furthering our precision engineering capabilities and ensuring that the Air LiHa offers low maintenance, high performance operation.”

The Air LiHa Assembly Team (back row, left to right: Sven Schmidt, Claudio Monti, Firat Sert, Kaspar Städeli, Thorsten Jörg, Jörg Egli) together with Project Leader Urs Schumacher (front left) and Product Manager Ralf Masantschek (front right)

Fredy Hottinger, Head of Assembly in Männedorf, added: “The transfer of any new product from development to production is always a critical step in terms of product quality. We have worked more closely with R&D than ever before on the Air LiHa – it has been a real

team effort – and we are proud of the quality and performance of this exciting new offering, and the benefits it will bring to our customers.” To find out more on Tecan’s Air LiHa, visit www.tecan.com/airliha

Corporate NEWS TECAN JOURNAL 1/2013

Historic Boston plays host to the fifth Tecan Symposium

Boston 2012 fifth Tecan

Mass spectrometry (MS) experts gathered in the historic city of Boston for the fifth annual Tecan Symposium, exploring how this technology is becoming a critical tool for a wide variety of applications. Building on the previous year’s success in China, the 2012 Tecan Symposium ventured across the Pacific to Boston, USA, where this year’s topic – Mass spectrometry: the expanding role in life sciences and diagnostics – gave delegates an insight into some exciting opportunities and new developments in this fast-moving field. Marc Feiglin, Chief Technology Officer for life sciences at Tecan, opened proceedings, welcoming delegates and commenting on the multitude of seemingly unrelated topics linked by MS. The day’s presentations began with John Brennan from McMaster University, Canada, who described the use of electrospray and MALDI-MSMS for biological screening of small molecule mixtures, including an intriguing overview of a new assay involving magnetic ‘fishing’ to identify modulators of protein-protein interactions. Robert Moritz from the Institute for Systems

Biology, USA, followed, with a discussion on the development of the complete human PeptideAtlas and SRMAtlas, a valuable publicly available resource. Moving on to the use of MS in medical research, Mitsutoshi Setou from the Hamamatsu University School of Medicine in Japan explained how lipids – which are traditionally quite hard to visualize – can be investigated using high resolution imaging MS, aiding studies into conditions such as atherosclerosis and Alzheimer’s disease. Sabine Becker from the Juelich Research Centre in Germany drew the session to a close with a fascinating talk about BrainMet and the use of MS to investigate the role of metals in neurodegenerative diseases. Gary Van Berkel from the Oak Ridge National Laboratory, USA, began the afternoon session, giving delegates an insight into the analytical opportunities generated by

Symposium

combining the high spatial resolution of laser desorption/ablation with liquid extractionbased surface sampling probes. Judy Stone from the TPMG Kaiser Regional Laboratory – Northern California, USA, explained how MS is benefitting clinical laboratory applications, and described how an automated LC-MSMS assay for 25-hydroxy vitamin D is enabling more than 1,500 samples to be analyzed each day. The results of a Friday night chance experiment were discussed by Burak Eral from the Massachusetts Institute of Technology, USA, who described the concept of eMALDI, which uses electrowetting to suppress the ‘coffee stain’ effect. He was followed by Feixia Chu from the University of New Hampshire, USA, who used MS-based proteomics to investigate how proteins survive in amber samples and fossils. Paul Tempst from the Memorial Sloan-Kettering Cancer Center, USA, rounded off the day by raising the dire rate of progress in disease

Robert Moritz discussed the development of the complete human PeptideAtlas and SRMAtlas Symposium speakers and chairpersons gather for a photo shoot

Corporate News TECAN JOURNAL 1/2013

Delegates discovered Boston’s historic landmarks during a tour of the Freedom Trail

biomarker discovery and the need to look at the activity, and not just the presence or amount, of proteins. Day two of the Symposium put the spotlight on the use of MS as a routine diagnostic tool, where robust, accurate assays are vital, due to the direct impact results can have on clinical decisions and peoples’ lives. Nigel Clarke from Quest Diagnostics, USA, discussed how patients may undergo unnecessary treatment as a result of false positive results obtained using immunoassays when, in many cases, this could be avoided by the development and use of more specific LC-MSMS assays. The next speaker, Michael Vogeser from the University of Munich, Germany, focused on the need for full automation of the entire clinical LC-MSMS protocol, explaining the pitfalls associated with MS analysis and pre- and post-MS sample processing, and concluding that automation is essential to further improve the reliability of these applications. Andy Hoofnagle from the University of Washington, USA, went one step further, suggesting that, while immunoassays have been – and remain – a useful tool, companies needed to work in partnership to advance the development of fully automated MS-based clinical assays. MS-based neonatal screening for treatable metabolic conditions was then discussed by Xuefan Gu from China’s Xin Hua Hospital, who commented that ‘prevention is better than cure – better to build a fence at the top of the cliff than a hospital at the bottom’. The concluding speaker of this session, Zoltan Takats from Imperial College London, UK, had the audience on the edge of their seats as he described how REIMS (rapid evaporative ionization MS) technology

is enabling surgeons to obtain real-time information about tumors during cancer surgery, allowing much more accurate removal of malignant tissue. The final session of the Symposium was opened by Jeff Hurst from The Hershey Company, USA, who took delegates back to 600 BC (before chocolate), describing how MS techniques have enabled the identification of chocolate residues on ceramic vessels recovered from archeological sites. Mark Libardoni from the Southwest Research Institute, USA, then took the audience on a journey into space, explaining the value of MS in space science applications, before Mehdi Moini from the Smithsonian Institution, USA, drew the Symposium back to Earth with a talk on the successful application of ultrafast capillary electrophoresis MS to dating museum samples. Networking opportunities and social events – including a tour of the Freedom Trail and an evening dinner in Boston Public Library – allowed delegates to continue discussions in an informal atmosphere, before heading home to contemplate this vast array of MS applications and the success of the 2012 Tecan Symposium.

“Prevention is better than cure – better to build a fence at the top of the cliff than a hospital at the bottom.”

To find out more about the 2012 Tecan Symposium, visit www.tecan.com/symposium2012

Left to right: Denis Coulet, Marina Lomtatidze, Natalia Mudrak and Mark Hozza enjoy an informal discussion over a glass of wine

CORPORATE NEWS TECAN JOURNAL 1/2013

Introducing David Martyr Just a few weeks into his new role, Tecan Group CEO Dr David Martyr took some time out of his busy schedule to talk about his previous roles and experience, and look ahead to a challenging and exciting future with Tecan.

"I have developed a huge amount of respect for the Company. Tecan has a much larger market impact and reputation than most companies of its size."

Please tell us a little about your background in technology and life sciences. I actually studied naval architecture and ship building at university, which is a bit of a strange route into life sciences, but I have always been fascinated by all forms of technology and engineering. After a year working as a naval architect – there are a few ships still afloat which I had a hand in designing – I went back to university to do my PhD, and took the opportunity to change disciplines slightly, looking at the applications of laser technology in ship building and heavy industry. This eventually led me into the laser industry, where I found that I really enjoyed interacting with customers and learning about their applications. I have

switched fields several times since then, but throughout my career I have always been drawn to learning about new applications and technologies. My background in lasers and optical technologies eventually led me into life sciences, when I joined Leica’s confocal laser microscopy and image analysis business in 1998. I did not have a huge amount of experience in the life sciences sector at that time, but was fascinated by the sheer breadth of technologies and applications, which ranged from cutting-edge microscopy hardware for academic centers to genotyping software development for clinical applications. This was also my first exposure

CORPORATE NEWS TECAN JOURNAL 1/2013

to Tecan, as part of a collaborative project to develop an early version of the GenePaint™ software. What was your initial impression of Tecan? I have developed a huge amount of respect for the Company. Tecan has a much larger market impact and reputation than most companies of its size, and that was something that interested me from the start. I have had extensive dealings with Tecan’s OEM business over several years – Leica Biosystems is one of the biggest customers of Tecan Cavro® brand components – and I have always found it a fascinating company. Tecan has continually focused on innovation and has built a strong reputation for quality, both attributes that are very important and attractive to me personally. I have followed the Company’s progress with interest, knowing that I would be very proud and pleased to perhaps one day be involved in its future development. You’ve had many successes in your previous roles. How do they resonate with the challenges you see at Tecan? The market has changed significantly over the last few years, becoming more and more application focused. Where companies may previously have been structured around separate, semi-autonomous geographical territories, there is now a need for a more integrated, application-driven approach. Customers are looking for far more than a straightforward purchase of capital equipment; it has become a consultative selling process, with manufacturers now expected to provide exceptional hands-on technical and applications support. This requires a change of business culture similar to that which my team and I achieved at Leica, restructuring companies not just to reduce costs, but also to break down internal barriers. Combining this with an open and trusting environment allows people to take

ownership and responsibility of projects, safe in the knowledge that senior management will not seek to blame individuals if something goes wrong, and enables a company to better focus on delivering on its brand values. Finally, what are your short- and long-term plans for the Company? Tecan has always been a pioneer in laboratory automation, but the market does not stand still, and we need to reaffirm our reputation as a leading supplier of best-in-class equipment. We already have a good sense of the opportunities within the market, how we can develop and what products and technologies we need to focus on and expand into. My challenge is to raise the organic growth of the business by bringing successful new products to the market in a repeatable, consistent way. It is always difficult to put structured management processes in place without killing the innovative spirit within a company, and rapid growth can become a very tiring environment, as a large part of any success is down to hard work. However, when you start to see the results of that effort, it is very motivating, and I hope to create new momentum – a wave of innovation – which will challenge and inspire our staff. Our ability to work in partnership with other companies will also be key to this. Open innovation, collaborating with other leading technology companies, will help to drive forward development of new technologies and provide new opportunities. Finally, a lot of enthusiastic and talented individuals have joined Tecan in recent months and years, and we need to make sure that we harness their skills to complement the Company’s established team, striking out ahead to develop strong technologies and innovative products which will make Tecan the obvious choice for cutting-edge laboratory automation solutions.

Readers TECAN JOURNAL 1/2013

A watchful eye on biosecurity The Infinite® F500 is BioSentinel’s reader of choice for its high sensitivity botulinum neurotoxin assays. Intended for government laboratory and pharmaceutical screening applications, BioSentinel now recommends the Infinite F500 to all its customers, thanks to the reader’s combination of advanced features and competitive pricing.

BioSentinel Pharmaceuticals, based in Madison, Wisconsin, USA, was founded in 2007 to develop and commercialize rapid, high sensitivity assays for the detection of the botulinum neurotoxins (BoNT). These toxins block neurotransmitter release at the presynaptic terminal of nerves, and are commonly used in the medical and cosmetic

sectors, as well as having bioweapon potential. The Company offers a range of both biochemical and cell-based assays, which are used by the US Department of Defense for detection of bioterrorism agents, and by large pharmaceutical manufacturers for potency testing of BoNT-based drug products and substances.

Dr Ward Tucker, Director of Research and Development at BioSentinel, explained the Company’s approach: “Our assay technology is designed to measure the proteolytic activity of BoNT down to the low picomolar or femtomolar range. Based on fluorescence detection, our assay uses a conjugated cyan fluorescent protein/yellow fluorescent protein (CFP/YFP) substrate which is cleaved by BoNT. In the absence of any toxin, excitation of the CFP results in fluorescence from the YFP due to FRET between the two domains. If BoNT is present in the sample, it cleaves the protein conjugate, leading to an increase in CFP fluorescence and a corresponding decrease in YFP fluorescence. By measuring the ratio between the CFP and YFP, we are able to accurately quantify BoNT activity. This mechanism of detection is highly sensitive, and the assay has been developed as both a biochemical assay, BoTest™, and a cell-based test, BoCell™.” “We initially developed the assay kits with a high end, monochromator-based reader, but we wanted a more cost-effective solution that we could recommend to customers using the assay for routine screening applications. Although the BoTest has fairly straightforward detection needs, the BoCell kit, like any adherent cell-based assay, has more complex testing requirements to ensure robust and consistent data. One of our primary requirements was a reader that could measure at multiple user-defined sites within each well, allowing us to compensate for natural variations in cell density across the well, as well as avoiding the areas where cells may have been displaced by pipetting.”

The Infinite F500 offers a high performance, cost-effective solution for BoTest and BoCell assays

“We tested microplate readers from several manufacturers, and the Infinite F500 from Tecan was clearly the best match for our

Readers TECAN JOURNAL 1/2013

Excitation 435 nm CFP

Emission 470 nm

2.5 2.5 1.5 1.5 0.5 0.5 10-1 10-1

YFP

102 10 (pM) 10 [BoNT] 103 1

101

103 104

1h 2h 4h 18 h

104

[BoNT] (pM)

The BoTest assay allows detection of BoNT activity over time

Infinite series reader for our application. We were given the option to test multiple systems, which is very important, as we would not either purchase or recommend an instrument that we hadn’t had the opportunity to test in routine operation. Since purchasing our instrument it has been in daily use by multiple users, and has proven both straightforward and reliable to operate. Although we use our own bespoke software to perform the data analysis, the

Emission 526 nm

FRET

1h 2h 4h 18 h

+BoNT

Excitation 435 nm

instrument’s control software is certainly user-friendly; it is easy to set up and run protocols. The reader has also been reliable over the last couple of years, we’ve not had any issues in terms of functionality and it always performs exactly as we expect.” To find out more about Tecan’s Infinite F500, visit www.tecan.com/infinitef500 To learn more about BioSentinel, visit www.biosentinelpharma.com

FRET

Emission 526 nm

CFP

Emission 470 nm

BioSentinel’s FRET-based assay technology detects the proteolytic activity of BoNT using a cleavable CFP/YFR conjugate

YF P

“We are also very happy with the support we received from Tecan during the evaluation process. The Company’s local representative took the time to understand our exact requirements, and provided us with a very good overview of the benefits of each

BoNT/A BoNT/A emission ratio emission ratio (Em 526/Em 470) (Em 526/Em 470)

needs. The standard filter sets are well-suited to our assay, and it offers us the flexibility to optimize the number and location of reads within the well. The Z-optimization feature is also a bonus, as this automated function ensures that the maximum signal intensity is achieved, further improving the sensitivity of our assay at low BoNT concentrations. We wanted a reader that we could recommend to customers who did not already have fluorescence detection capabilities, so it had to be cost-effective, but the quality of the data achieved using the Infinite F500 was also superior to the competitors’ instruments for our assay kits. This, combined with Tecan’s excellent reputation and the fact that several other molecular diagnostic companies recommend Tecan readers, made the Infinite F500 the logical choice.”

food APPLICATION TECAN JOURNAL 1/2013

Baked to perfection Puratos is using a Freedom EVO® 200 workstation to aid the development of novel enzymes for use in the baking industry. By automating enzyme screening and characterization, the Company is able to increase throughput and productivity, as well as improving quality control testing. Puratos is an international manufacturer of innovative products and raw materials for the bakery, patisserie and chocolate sectors. The Company’s bakery division develops a wide range of improvers which bakeries can use to alter the flavor, texture and shelf life of their products. The Enzymes Business

Unit, based at the Company’s facility in Andenne, Belgium, is responsible for the development of novel enzymes for inclusion in these baking ingredients, and has seen a steady growth in its workload since it was created in 1994. Valérie Dorgeo, Laboratory Supervisor for the Enzymes Business Unit, explained: “Our laboratory is constantly looking for new or enhanced enzymes which can improve or accelerate our customers’ baking processes. We start by screening new enzymes or by creating a variety of genetic variants of the enzyme of interest, then use in vitro activity assays to characterize these enzymes. Since

food APPLICATION TECAN JOURNAL 1/2013

David Denis and Valérie Dorgeo alongside the Freedom EVO 200 workstation

we were first established, the laboratory has seen a significant increase in both the number of projects we perform, and the number of samples per assay. To deal with this, we began investing in laboratory automation several years ago and have semi-automated several of our methods. However, even semi-automated protocols are prone to human error, and so we investigated the possibility of using a fully automated workstation to help us improve process security, as well as further increasing our throughput.” “We looked at a number of options, and the Freedom EVO workstation from Tecan offered the most flexibility in terms of the number of different protocols we could run, as well as being a very robust instrument. It is also very quiet, which has to be a consideration for any piece of laboratory equipment which is likely to be in constant use, and has a number of built-in security features to simplify day-to-day operation. Tecan was very helpful in terms of understanding our needs, and arranged for us to run one of our protocols on a demonstration instrument. This gave us complete confidence that our workflow could be automated in the way that we wanted.” Valérie continued: “Our Freedom EVO 200 platform is configured to be as versatile as possible, allowing us to continuously switch between assays. The instrument

is equipped with both eight-channel LiHa (Liquid Handling) Arm and MCA 96 (MultiChannel Arm™) pipetting options, as well as a RoMa (Robotic Manipulator) Arm for microplate handling, offering true parallel tasking. It also has a number of other devices integrated onto the deck of the instrument to allow on-board assay processing, including heated and shaking incubators, a thermal cycler and an Infinite® M200 PRO multimode reader equipped with a NanoQuant Plate™. Although there are currently several small steps which are performed manually – we do not yet have an integrated plate sealer or centrifuge – our intention is to completely automate around 10 of our assay protocols.” The increased process security offered by the Freedom EVO is also helping the Enzymes Business Unit to streamline its quality control (QC) activities. David Denis, QC operator, explained: “Because manufacturing is located on the same site as the enzymes development department, we have been able to take advantage of the reproducibility and precision offered by the Freedom EVO for QC testing. The system’s in-built process security features and PosID™ sample tracking are well suited to this application, providing us with additional guarantees that our products meet the stringent specifications of our rigorous QC program, as well as accelerating the overall speed of the QC process.”

Valérie concluded: “Since taking delivery of our Freedom EVO workstation, we have had very good support from our local Tecan representative, who has assisted us in transferring existing manual protocols to the workstation platform, as well as providing on-site training for several users. Although we are still in the validation phase, we are totally confident that we will be able to switch many of our current assays to the Freedom EVO, improving the quality of our data and streamlining our laboratory workflow.” To find out more on Tecan’s food applications, visit www.tecan.com/food To learn more about Puratos, go to www.puratos.com

Drug Discovery TECAN JOURNAL 1/2013

Speeding up drug discovery Production of dose-response plates and cherry-picking are essential steps in early drug discovery to enable testing of many thousands of compounds. Researchers at Novartis are benefitting from the automated plate generation capabilities of two customized systems based on Freedom EVO® 200 platforms, each equipped with an MCA 384, significantly increasing sample throughput to over 10,000 compounds a day.

“ The MCA 384’s adaptor plate concept gave us the flexibility to switch between 96- and 384-well formats.”

Yannick Gautier with the DRP system

The Novartis Institutes for BioMedical Research (NIBR) screen hundreds of thousands of compounds for subsequent lead optimization in early stage drug discovery. To meet the demands of today’s research, its Compound Management Group (CMG) in Basel, Switzerland, has embarked on a Compound Bank initiative to replace its existing hardware infrastructure, collaborating with Tecan to develop an automated system to deliver customized dose-response plates (DRPs). Frank Hoehn, a laboratory head within CMG, explained: “In 2009, we decided to replace the liquid handling module of our old Solution Archives (SolArs) system. We contacted a number of vendors, and Tecan offered us the opportunity to test a Freedom EVO platform with a MultiChannel Arm™ (MCA). The platform’s small footprint and high efficiency made the Tecan system an attractive choice, and it also offered the best technical fit for our needs in terms of quality, throughput, variability and software capabilities. The Company’s attitude

also played a major part in our decision; the Tecan Integration Group (TIG) showed genuine interest and fully engaged with us.” The CMG purchased two identical DRP systems, designed for automated plating of compounds from Matrix storage tube racks into 96-, 384- and 1,536-well destination plates. Fully integrated with the Company’s LIMS, each Freedom EVO platform is equipped with MCA 384, Liquid Handling (LiHa) and Robotic Manipulator (RoMa) Arms, as well as an integrated Cytomat® 24 hotel and docking station (Thermo Scientific), a PlateLoc® Thermal Microplate Sealer (Agilent), a Multidrop® Combi nL (Thermo Scientific) for bulk filling, an argon bath, a customized 96 piercing head, and a variety of MCA 384 adaptor plates. Research investigator Caroline Engeloch commented: “The MCA 384’s adaptor plate concept gave us the flexibility to use both disposable tips and washable fixed tips, as well as to switch between 96- and 384-well formats, and we worked with the TIG team to design customized adaptors that can pick two or three rows or columns of tips simultaneously, increasing the throughput of the system. We also needed to be able to use fixed tips on the MCA to pipette directly from Matrix tubes in a 96-well format, without having to uncap and recap the tubes. This required a bespoke piercing head to transfer the entire contents of a 96-tube rack into a 96- or 384-well plate, and Tecan was the only vendor to offer a solution with the flexibility we were asking for.” Caroline continued: “After delivery of the system, we performed site acceptance testing, optimization and validation to ensure that the platform met all our requirements. With no specification to start from for the customized piercing head, there was a lot of

Drug Discovery TECAN JOURNAL 1/2013

testing to be done, including quality control, reliability, variability and durability of the tips. The system also combines many different dispensing technologies to provide efficient generation of high quality plates and we collaborated closely with Tecan to optimize pipetting and tip washing conditions for our needs, minimizing the volume of wash solution required and reducing cross-contamination.” “The Freedom EVO-based platforms are far more flexible than our old system. The userfriendly software allows us to change and adapt the workflow to our needs, and we can implement new dose-response plate layouts much more quickly. We are also happy with the support we receive and the ease with which we can introduce new protocols; we occasionally need to contact Tecan, but we can implement most of the changes ourselves. This gives us confidence that the system is reliable and working as it should, increasing our productivity and ensuring we can meet our customers’ deadlines.”

A 3D representation of the Novartis DRP system

Frank concluded: “Our system is now operational and in routine use, and during the test phase we were able to process 5,000 hits per DRP module – 10,000 per day – increasing our throughput five- to six-fold compared to our old SolArs system. Our goal now is to have the same DRP system on both sides of the Atlantic, helping us to more closely align our processes, and so a similar platform is currently being planned for our site in Cambridge, USA.” To find out more on Tecan’s customized solutions, visit www.tecan.com/tig To find out more about the NIBR, visit www.nibr.com

The DRP project team. Left to right: Caroline Engeloch, Daniel Baeschlin, Ingrid Beuttenmueller, Frank Hoehn, Thomas Steiner and Yannick Gautier

Microarrays TECAN JOURNAL 1/2013

Developing a proteomic fingerprint for pancreatic cancer Scientists at the German Cancer Research Center are using an HS 4800™ Pro microarray hybridization station and a PowerScanner™ to investigate pancreatic cancers. By combining these automated solutions with in-house developed antibody microarrays, the Center is able to study pancreatic cancer at a protein level, helping to better understand the basic biology of the disease and develop new screening strategies for early detection.

Pancreatic cancer has a very high mortality rate as symptoms can go undetected for many years before diagnosis, limiting the possibility for treatment. To help alleviate this problem and improve survival rates, the Deutsches Krebsforschungszentrum (DKFZ, the German Cancer Research Center) is using protein microarray techniques to help develop early diagnostic and prognostic markers. Dr Jörg Hoheisel from the DKFZ’s Division of Functional Genome Analysis, explained: “Our group is focused on the development of new techniques which can be applied to routine diagnostic applications or can help to understand the complex biology of pancreatic cancer. We have had a strong interest in microarray technologies for a long time and moved into the proteomics field about a decade ago, developing a number of peptide, protein and antibody microarrays to look at various aspects of the disease.” Dr Christoph Schröder continued: “One of our major projects has been the development of an antibody microarray targeting over 740 proteins of interest in pancreatic cancer1. The target proteins are generally either strongly up- or down-regulated in tumor cells, and this microarray is designed to allow the investigation of the key cellular pathways involved in cancer, as well as to help identify biomarkers of diagnostic or prognostic value. Throughput and reproducibility are crucial to enable the generation of meaningful data in this type of application. We have been using an automated microarraying platform and a system with automatic mixing during incubation for some time, and have been able to achieve very good results. However, all washing and drying steps were performed manually, and so we looked at the possibility of further automation of our workflow to increase throughput and further improve reproducibility.” “We trialed hybridization stations from various manufacturers, and the Tecan HS 4800 Pro provided the best solution for our needs,” Christoph said. “Automation of the washing and drying steps offered the

Microarrays TECAN JOURNAL 1/2013 Signal-to-noise (S/N) ratios for multiple replicates of antibody microarrays representing 741 different cancer-related proteins. Incubations were performed in quadriPERM® chambers:

200.0

(1) without mixing

Signal – Background – Ratio

100.0

(2) with mixing on a standard lab shaker

50.0

(3) with an alternative commercial system for microarray incubation, in all three cases using manual washing and drying steps, or

20.0

(4) fully automated using the HS 4800 Pro.

10.0

Red bars depict the S/N ratios for the red fluorescence channel, and green bars correspond to the S/N ratios for the green channel

5.0 2.0 1.0

robustness and low variability we needed, as well as improving throughput. It also significantly improved the signal-to-noise ratio for our arrays, allowing us to reduce the incubation time from 16 hours to just 1-3 hours without compromising the quality of results, which increased our throughput even more.” “As we mainly work with clinical samples – both from hospitals here in Germany and as part of larger European projects – the sample volume required for analysis was another important consideration. Only very limited amounts of biological material are available, and samples cannot usually be replaced if an experiment fails. The HS 4800 Pro combines excellent reliability with very good reproducibility; we get the same data from any given sample, even if it is run on different days. As our throughput was increasing rapidly, we also looked at replacing our old microarray scanner. Again, we looked at instruments from several manufacturers, and felt that Tecan’s PowerScanner was the best fit for our laboratory. The system is intuitive to use, and its robust design gives you confidence in the results.”

HS4800 Pro (4)

other instrument (3)

standard shaker (2)

no mixing (1)

0.5

Christoph concluded: “This combination of instruments offers excellent reproducibility, and we are in the process of publishing several studies based on the data we have generated. We have also recently begun a collaboration based on this technology with a group at McGill University in Montreal, Canada, using the same instruments, microarrays and protocols in both laboratories to extend the use of the antibody array platform. Overall, we have been very pleased with the performance of the Tecan instruments. The biomedical results we have achieved have led to the creation of a spin-off company focusing on the development of diagnostic tests and custom services based on antibody microarray screenings.” 1. Schröder, C et al. Dual-color proteomic profiling of complex samples with a microarray of 810 cancer-related antibodies. Mol Cell Proteomics, 2010, 9, 1271-80. To find out more about Tecan’s HS 4800 Pro and PowerScanner, visit www.tecan.com/microarray For more information on the DKFZ’s Division of Functional Genome Analysis, visit www.dkfz.de/funct_genome

Dual-color image of an antibody microarray incubated with two different plasma samples

COMPONENTS TECAN JOURNAL 1/2013

Made-to-measure robotics LCTech GmbH has a long association with Tecan Systems, and has been using its Tecan Cavro® components for many years. The Company’s latest sample preparation workstation, the FREESTYLE, uses a Cavro Omni Robot and Cavro XLP Pump to provide automated preparation of foods, feeds and environmental samples. LCTech, based in Dorfen, Germany, has been developing and supplying products and methods for sample preparation and analysis since 1998. Focusing on trace contaminant testing of foodstuffs and animal feeds – as well as quality control testing for the pharmaceutical industry – the Company’s in-house developed product range includes both semi and fully automated systems designed to simplify preparation of samples for analysis. Dr Uwe Aulwurm, a Managing Director of LCTech, explained: “We have had a long relationship with Tecan, and have always enjoyed good support from our local representative. Our latest system, the FREESTYLE, is designed for laboratories preparing complex samples for quality control testing and contamination screening. To meet the varied workflow demands of our customers, we wanted a modular solution that combined a general purpose liquid handling system with a range of different processing devices, including options for solid phase extraction, gel permeation

chromatography and evaporation. We weren’t looking to reinvent the wheel by developing our own XYZ robotic system. The Cavro Omni Robot was perfectly suited to our needs, providing a very robust instrument which could be easily modified to access individual processing modules, as well as for planned future developments.”

“ We have had a long relationship with Tecan, and have always enjoyed good support from our local representative.”

“Liquid handling is performed by a single Cavro XLP Pump equipped with a 10 ml syringe for all three current applications. This pump is well matched to the needs of our customers in terms of liquid handling performance and long-term stability, and uses Cavro ceramic valves – either 9- or 12-port configurations – to ensure solvent compatibility. This set-up provides very consistent operation and maximum flexibility for our customers’ applications, and the seamless firmware and software compatibility with the Cavro Omni Robot made it the logical choice.”

closely match the needs of our customers, which can vary significantly from day to day. The platform is already in routine operation with several customers, and they all appreciate the different options offered by this compact and easy-to-use XYZ robotic system. Our cooperation with Tecan has been vital to achieving this flexibility, and we look forward to working with the Tecan team to develop further applications for the FREESTYLE system.”

“Use of Tecan Cavro components has allowed us to develop the FREESTYLE system to

To learn more about LCTech, go to www.lctech.de

Dr Uwe Aulwurm (left) and Michael Baumann (right), the managing directors of LCTech, with the FREESTYLE system

To find out more on Tecan’s Cavro components, visit www.tecan.com/components

The FREESTYLE can handle plastic or glass SPE cartridges up to 15 ml

Liquid Handling TECAN JOURNAL 1/2013

More than skin deep Symrise is using an HP D300 Digital Dispenser to improve throughput and reproducibility in its cell and molecular biology laboratories, offering more reproducible results for cosmetic active ingredient testing. Symrise AG, based in Holzminden, Germany, is the world’s fourth largest manufacturer of fragrances and flavors. The Company’s Scent and Care Division develops, produces and markets a wide range of fragrances and cosmetic active ingredients, specializing in innovative substances and technologies which confer efficiency or other additional benefits to its customers’ products, such as skin toning, multifunctional antimicrobial activity and sun protection, as well as active botanicals and products for sensitive skin. As part of the Company’s rigorous testing and validation program, the Innovation Life Essentials group performs a broad spectrum of cell-based assays, using skin-derived keratinocytes, melanocytes, fibroblasts and adipocytes to identify the most effective and safe compounds for inclusion in cosmetic products. Imke Meyer, Manager Cell Biology, at Symrise, explained: “We are predominantly a flavor and fragrance company, but the activities of our Life Essentials team are well recognized as the major differentiator within our global industry. We work with a broad

range of cutaneous and subcutaneous cell types, developing cell-based assays to identify and characterize novel active ingredients for inclusion in our customers’ cosmetic products, primarily relying on low or medium throughput ‘intelligent’ screening strategies, predominantly using manual techniques.” “We became interested in the HP D300 Digital Dispenser after seeing the system demonstrated at an exhibition in Hamburg. Frequently, the compounds involved in our screening trials are in limited supply, and we could clearly see the potential of this instrument to eliminate the need for the wasteful serial dilutions. Direct digital dispensing into the assay plate is also faster and more accurate than manual pipetting, helping to improve the reproducibility of results while improving our throughput.” “We purchased our system in early 2012, and have already seen the benefits for a variety of assays. For example, in a recent cytotoxicity trial we were able to switch from performing an IC50 determination based on four different

concentrations in triplicate, to a single series of measurements at eight different concentrations, thanks to the improved reproducibility offered by direct titration. This not only results in better data, allowing more accurate calculation of exact inhibitory concentrations, it also has significant cost benefits in terms of the quantity of cells, reagents and consumables required per assay. Similarly, we have been able to reduce the number of replicates required for several of our other assays, and the system’s intuitive software is certainly encouraging us to use it wherever possible.” To find out more about the HP D300, visit www.tecan.com/digitaltitration To learn more about Symrise, visit www.symrise.com

“ ...we could clearly see the potential of this instrument to eliminate the need for the wasteful serial dilutions.” Left to right: Mirjam Knupfer, Julia Betke and Ann-Christin Weseloh using the HP D300 Digital Dispenser

Readers TECAN JOURNAL 1/2013

Too gracious a host Legionnaires’ disease, a potentially fatal infection caused by Legionella pneumophila has recently been in the headlines due to a number of serious outbreaks around the world. Scientists at the University of Toronto are using Tecan’s Infinite® M200 PRO and Gas Control Module to study host-pathogen interactions for Legionella bacteria. Legionnaires' disease is an uncommon form of pneumonia which is fatal in 10-15 % of the general population. Although the bacteria are widely distributed in the environment where they can live in all types of water, they only become a risk to health in conditions that allow Legionella to grow rapidly, such as in poorly designed or maintained water systems.

Researchers at the Ensminger Lab, part of the Department of Molecular Genetics at the University of Toronto, Canada, are investigating host-pathogen interactions, specifically for Legionella pneumophila, using cutting-edge genomics, experimental evolution and high throughput robotic solutions. Assistant Professor Alex Ensminger

The Ensminger Lab team (left to right): Alex Ensminger, Chitong Rao, Carly Weiss and Amy Chung

discussed the laboratory’s approach to studying this potentially deadly pathogen: “Legionella pneumophila normally grows inside host cells; it is not thought to grow in the water as such, but rather in amoebae and protozoa present in the water. The bacteria are basically professional pathogens of amoebae and amateur pathogens of human

Readers TECAN JOURNAL 1/2013

cells; the human lung is an evolutionary dead end where the bacteria cause disease. We need to understand how these pathogens interact with both their natural hosts, and accidental hosts – more specifically, patients’ lungs – to learn how growth in those two situations is similar but, equally importantly, different. This knowledge is critical to understanding how these bacteria persist in different environments, growing to high levels in cooling towers, for example, and how this offers them the opportunity to cause human disease.” The Group’s starting point was to experimentally evolve the bacteria, passaging them repeatedly in specific types of amoebae or macrophages, and observing any changes to see if they became more specialized for different hosts. A key part of this procedure is to monitor bacterial growth under certain conditions, which is a very labor intensive process, involving plating out, counting colony forming units, and comparing different strains with each other. Alex soon identified an alternative: “I had seen recent publications describing the use of the Infinite M200 PRO to measure CO2-independent growth of Legionella using a green fluorescent protein (GFP) marker. As soon as I read that the instrument was also available with a Gas Control Module (GCM™), I was sold on buying one. I realized that I could put the bacteria in host cells, place them in the plate reader and leave them there for three or four days, watching their growth. By measuring every 20 minutes or so, we have seen growth curve shapes that disappear if readings are taken less frequently, even hourly. These shapes match what we might predict from cell biology, and allow more subtle distinctions between phenotypes than was previously possible.

Growth is observed for 14 hours, followed by a pause of about four hours before it begins again, which reflects what we know, that an initial round of infection takes 14 hours, and then the bacteria lyze out of that first set of host cells and cause further infection.” “We use both fluorescence and luminescence measurement in our experiments and, with the high throughput that the Infinite M200 PRO offers, we are able to look at the same strain under a variety of conditions simultaneously, minimizing day-to-day variations and giving us extra confidence in our results. We soon realized that we needed another reader, and it made sense to integrate this with a Freedom EVO® 100 platform to give us even more flexibility and the capacity to increase throughput going forward. An incubator stack maintains 96-well plates at a set temperature, and every 20 minutes the Robotic Manipulator (RoMa) Arm transfers a plate to the reader. After dark adapting to prevent autoluminescence background signals, a luminescence measurement is taken. Fluorescence measurement is even simpler.” Alex and his team have widened their studies as they have become more familiar with their Tecan instruments: “We have some evidence that nutritional requirements influence the host range of the bacteria and how they are recognized by the host cell. To investigate this, we are using the Freedom EVO to prepare batches of 96-well plates where each well is lacking a different nutrient, and can seal and freeze them for future use. Along with our colleagues at Public Health Ontario, we are also using this technology to look at clinical and environmental strains that have been collected over the last 30 years, trying to identify phenotypic and genetic diversity

and determine whether strains from specific outbreaks are more specially adapted to different hosts. We are moving towards a situation where we will be able to establish the specific isolate or type of bacterium causing an outbreak and make predictions regarding its host range and virulence. The aim is to see if there are conditions which select for more virulent bacteria that are more likely to cause human disease. The idea is that there may be specific natural hosts which select for more virulence, or perhaps if a specific type of protozoan or amoeba is present in the water system, that could act as a biomarker for the system being at increased risk for selecting more virulent bacteria. Increased throughput and the capability to do phenotypic screening in a huge number of host cells simultaneously is the thing that’s making this possible.” To find out more about the Infinite M200 PRO and GCM, visit www.tecan.com/infinite200pro To find out more about the Ensminger Lab, visit individual.utoronto.ca/ensminger_lab

Proteomics TECAN JOURNAL 1/2013

Feeding the SRMAtlas project A Freedom EVO® 150 workstation is being used to great effect by the proteomics group at the Institute for Systems Biology (ISB), Seattle, to generate samples for analysis by mass spectrometry, including for the world-renowned Selected Reaction Monitoring Atlas (SRMAtlas) project. The system is equipped with a variety of modules to meet the requirements of a range of different projects, and provides the reproducibility and throughput to enable these studies to be run entirely in house. The multidisciplinary ISB in Seattle, USA, was established in 2000, and is involved in collaborative studies on model organisms – such as halobacterium and yeast – monitoring the genetic, translational and proteomic effects of biological perturbations on the functioning of systems within the organism. The proteomics department at the ISB, led by Dr Robert Moritz, comprises a team of investigators and computational biologists working together on multiple projects and collaborations. Research Associate Doug Spicer joined the team to run automation in the department, and took over a Freedom EVO 150 workstation to develop and routinely run automated protocols for four separate projects. These include automated methods for immunoprecipitations, phosphokinase enrichment and in-gel protein digests with trypsin, plus assay development for the human SRMAtlas, the collaborative project between ISB and the Swiss Federal Institute of Technology (ETH) Zürich to develop a resource for quantification of all 20,000 human proteins for deployment in human tissues, cell lines and blood. Doug explained: “The system was purchased two years ago, originally to perform reproducible plasma glycocapture to generate samples for mass spectrometry. The platform is capable of rapidly processing hundreds of crude peptide samples to create multiplexed assays, generating sufficient samples in one day to ‘feed’ three mass spectrometers over a three to four day period, producing data for the SRMAtlas project. It is an excellent, robust and flexible system fitted with a range of modules to meet the varying specific needs of each of our projects.”

The ISB team with the Freedom EVO. Left to right: Meg Kapousouz, Doug Spicer, Sarah Li

The ISB system can perform both singleand multi-channel pipetting using an eight-channel Liquid Handling Arm, and the Robotic Manipulator Arm interfaces with a variety of modules to maximize flexibility. These include a Te-Shake™ module for immunoprecipitation and phosphokinase enrichment protocols, and a Te-VacS™ vacuum manifold for in-gel digests and the SRMAtlas project. Doug added: “Separation of the fragments from in-gel protein tryptic digests is performed using a vacuum manifold plate with micro-holes drilled in the bottom of its wells. To avoid occlusion of these holes – which would prevent the reagents from draining through the plate into the vacuum manifold – hexagonal beads are placed in the wells of the microplate to act as

a filter and the gel fragments are placed on top. This allows the plates to drain correctly at a lower vacuum manifold pressure, and enables us to achieve good recovery of fragments from protein digests.” As differing protocols require either a static or shaking incubator, a third-party thermal mixer was integrated onto the workstation, with assistance from the Tecan applications team. The system’s software has a liquid class feature that helps to ensure effective, accurate handling of different liquid types – such as detergent, DMSO and acetonitrile – within the same protocol. He continued: “Having the ability to establish different ways of pipetting very

Proteomics TECAN JOURNAL 1/2013

different liquids has proven very helpful in overcoming the problems associated with handling these solutions precisely. Most significantly, although we are using fixed pipette tips, there are no problems with cross contamination and we are achieving high consistency and reproducibility between runs, which is essential in proteomics. Sample throughput and accuracy of pipetting are critical in the human SRMAtlas project, ensuring consistent representation of each peptide in one pool taken from all samples in one 96-well plate. These 96-well plates contain peptides characteristic of the original proteins, and over 1,700 96-well plates represent the whole of the human SRMAtlas, a compendium of the human proteome. Our throughput ranges from 20 to 24 plates daily, and the Freedom EVO’s volumetric accuracy makes consistent representation of each peptide in the pool possible – an advantage over manual protocols. This has enabled us to carry out the project entirely in house.”

Doug concluded: “I really appreciate the time and effort Tecan invests in developing its systems. We have a very good rapport with the application specialists and service engineers, and feel they really take on board the importance of refining the liquid handling features that are critical in most projects. If ever I find I can’t put together an appropriate automated protocol using my own experience, I know that Tecan will give me the help I need.” To find out more on Tecan's proteomics solutions, visit www.tecan.com/proteindigest To learn more about the Institute for Systems Biology, Seattle, USA, go to www.systemsbiology.org

“ Most significantly, although we are using fixed pipette tips, there are no problems with cross contamination.”

Developments at ISB – human SRMAtlas Synthetic ‘proteotypic’ peptide

Human proteins (from natural source or synthetic)

exity compl ion in reduct

(from cheap synthesis)

Develop human PeptideAtlas (from tryptic digests or synthetic peptides)

Develop human SRMAtlas (verified quantitative assays)

The human SRMAtlas process

Develop optimized transitions from PeptideAtlas

Microarrays TECAN JOURNAL 1/2013

Rewriting the proteomics handbook A suite of Tecan’s microarray products is ensuring that researchers at the Biodesign Institute in Arizona, USA, forge ahead with a range of functional proteomics projects looking at early biomarkers for several diseases.

The Virginia G. Piper Center for Personalized Diagnostics at the Biodesign Institute, Arizona State University, takes a multidisciplinary approach to looking at first line diagnostics, drawing on the expertise of engineers, physicists, molecular biologists and clinicians for a range of different diseases. Dr Joshua LaBaer, Director of the Center, explained: “We have a broad range of interests in our laboratory, and many of the biomarkers we are looking at are for cancers, for example, breast, ovarian and lung. Other projects, however, are targeted toward autoimmune diseases, like arthritis and diabetes, and, more recently, inflammatory bowel disease. Precision medicine underpins almost everything we do, and is changing the way that everyone views medical diagnostics.” “A key element of our approach is functional proteomics and we have adopted a completely unique technology for protein

microarrays,” Dr LaBaer continued. “Most groups who make protein arrays do so by developing high throughput platforms to purify proteins, and then spot the proteins onto the slides. However, this method doesn’t work well for purifying proteins, particularly with regard to yield. We prefer to print the gene for the proteins on the slide, and then add a cell-free extract that synthesizes the proteins in situ. The proteins are made literally an hour before we test them. Then, for personalized diagnostic applications, we probe these arrays with serum from patients, looking for responses to certain proteins indicative of disease.” The technique is pivotal to many of the projects within Dr LaBaer’s laboratory, and automation in the form of an HS 4800™ Pro automated hybridization station and two PowerScanner™ units has become essential to keep on top of the workload and produce

Some key members of the Center’s staff. Front row (left to right): Dr Josh LaBaer and Dr Ji Qiu. Back row (left to right): Mike Gaskin, Dr Mitch Magee and Alex Mendoza

consistent and reliable results. “These are our newest pieces of Tecan equipment and they are proving to be real workhorses in the laboratory. The PowerScanners are essential because we need to repeatedly scan our slides on a regular basis, and the optics are very good, giving us nice strong signals. A huge consideration for our work is having statistically valid sample sizes; does each study have enough statistical power to get the answers we need, especially taking into account that there is a huge amount of biological variation from person to person? For this reason, we make sure that we always have adequate study sizes and, as a consequence, we’re running many slides for many patients over and over again. This can be extremely tedious, and having reliable autoloaders on the PowerScanners has been extremely helpful. The accompanying software has also been very useful, enabling us to very quickly identify the features that have signals, measure those signals and then do all the different operations on them. We are now developing a full LIMS for the protein array platform, and the scanners will eventually be connected into that.” Historically, when the method was first developed, everything was done by hand, including all washing and incubation steps. “The problem in a high throughput setting is that human beings are simply not as precise. The HS 4800 Pro has been a really big advantage for us; it basically runs by itself overnight and has done wonders in terms of reproducibility from slide to slide. It allows us to produce these proteins in situ on the glass, wash them off, and even incubate them with samples, all in a single run, without any manual intervention at all.”

Microarrays TECAN JOURNAL 1/2013

Dr LaBaer has had Tecan instruments for more than ten years, including larger liquid handling workstations that were used to build a library of full length sequence clones for encoding human proteins. However, the team had a good look around at other microarray products on the market before making their choices. “In both cases, the devices were by far the best thing on the market. In the case of the PowerScanner, there was nothing out there with the same combination of good autoloading features, images and software. Other devices simply didn’t match the quality.” The same was true for the automated hybridization station, as Michael Gaskin, Center Manager and Automation Scientist, explained: “We tried a number of different systems, but this was the only one that worked well for us. There really was nothing that competed with its capabilities. Like all new things, it took a little time to develop the right protocol and, when we had the instrument for demonstration, Peter Herzer from Tecan helped us to work out how best to apply it to our workflow. Having it here and being able to test and work with it was really critical; if it wasn’t going to improve our pipeline, there was no point in buying it. The data that we had from that time is really what convinced us that this was the right tool for our laboratory, and it now has long waiting lists because it is in such high demand.” Dr LaBaer concluded: “When you’re trying to draw a conclusion about a clinical sample – about people – then the technology has to be spot on in terms of reproducibility. If not, your conclusions are going to be tainted by artefacts from the actual processing. With the HS 4800 Pro, we can do the same array

The Center’s PowerScanners provide reproducible, high throughput scanning

on two different days and still get the same answer. The reproducibility this instrument gives us has made all the difference.” To find out more on Tecan’s microarray solutions, visit www.tecan.com/microarray

Dr Fernanda Festa loading slides onto the HS 4800 Pro

To find out more about Dr LaBaer’s work at ASU, visit labs.biodesign.asu.edu/labaer To find out more about the plasmid collections created by the Center, visit dnasu.asu.edu

Readers TECAN JOURNAL 1/2013

Undergraduate studies enter a whole new world Tecan’s Infinite® M200 PRO multimode microplate readers are helping McMaster University to introduce chemical biology undergraduates to some of the most advanced techniques routinely used in today’s laboratories.

Chemical Biology undergraduates William Denk and Kevin Yin get to grips with the Infinite M200 PRO microplate reader

“ Using the Infinite microplate readers is a revelation for our students.”

In 2008, McMaster University in Hamilton, Canada, launched a new undergraduate program, Chemical Biology, with an emphasis on the development of a detailed understanding of the molecules governing biological phenomena. A series of integrated experiments, specifically designed to explore interactions between small organic molecules and large biochemical macromolecules, introduces the fundamental aspects of the subject and provides students with hands-on experience of sophisticated analytical techniques. John Brennan, Canada Research Chair in Bioanalytical Chemistry and Biointerfaces, explained: “When we established the Chemical Biology program, we were in the fortunate position of being able to start from a completely blank piece of paper, so we could do almost anything we wanted to. That gave us the opportunity to be innovative in the way we set up the course, rather than following a conventional structure.” “The first term of the second year includes an analytical course that teaches subjects including fluorescence, high throughput screening and plate-based assays. Although much of the practical work includes fairly conventional analytical techniques – such as titration, spectrophotometry and electrochemical measurements – we also wanted to introduce the students to the concept of high throughput bioassays. Therefore, in the second term, students begin to work with the Infinite M200 PRO microplate readers, equipped with absorbance, top and bottom reading fluorescence intensity and luminescence capabilities. Undergraduates learn the basics of plate reading, including the issues associated with using the outer wells, how to set up duplicates and where to place the controls, as well as the optimization of instrument settings such as broad or narrow bandwidths and top or bottom reading.

They realize that using cuvettes with three or four milliliters of reagent is not necessary; laboratories just don’t work that way anymore.” “We designed an 11 week program which is essentially one big experiment, as all the individual aspects are integrated together. The first few weeks are spent performing a natural product extraction, which is followed by the synthesis of a compound library where every student in the lab makes one variant of a particular compound. Over the next couple of weeks, they develop two different 96-well plate assays; a ligand binding assay and a kinetic assay. The ligand binding assay uses a dansyl glycine probe and BSA to introduce fluorescence measurement. If the dansyl glycine is displaced as a result of ligand binding, then a decrease in fluorescence intensity is observed; it’s a very straightforward assay. The kinetic assay is a colorimetric acetylcholinesterase assay which uses the Ellman reaction to produce a yellow color when DTNB – 5,5’-dithiobis-(2nitrobenzoic acid) – reacts with thiocholine. In this assay, the Infinite readers are used to measure absorbance. Developing these assays provides an opportunity to put theory into practice, for students to discover changes in intensity or wavelength for themselves, and to determine Km/Vmax and establish a Lineweaver-Burk plot. The remaining weeks are spent using these assays to screen both the natural products and synthetic compounds. The data is pooled and, if the initial extraction has been done correctly, they may be able to find a hit from the natural product and go on to do an IC50 determination as well. More importantly, the synthetic library is designed to allow students to perform a small QSAR (quantitative structure-activity relationship model) to work out which functional groups are actually important for enzyme inhibition.”

Readers TECAN JOURNAL 1/2013

John continued: “The third year lab builds on this knowledge, reinforcing the concept of making and screening molecules, and introducing cell-based assays. Students use an E. coli-based bioluminescent reporter, gaining experience in performing cellbased assays in a plate reader setting. We introduce additional aspects of fluorescence measurement – such as spectral shifts and how they can be used in assay development – and study the merits of monochromator- and filter-based readers.” “The Infinite M200 PRO was an obvious choice for our undergraduate laboratories. We already had a lot of Tecan equipment, including two Freedom EVO® liquid handling platforms, a Safire™ and an Ultra Evolution™,

as well as a couple of Infinite M1000s that are used as research instruments, and we particularly wanted monochromator-based instruments so that we could study spectra and look at using spectral shifts as a way of getting an assay to work. Students use the Infinite M200 PROs to begin with and, by the fourth year, will be using the Infinite M1000s with the additional features, such as fluorescence polarization. The software is almost identical, so it is a pretty simple transition from one instrument to the other. The instruments are also very robust, which is vital in an undergraduate environment.”

the first year performing basic analyses with large volumes of reagent, miniaturization – performing assays in 96-well plates with microliter reagent volumes – is a completely new experience, and they begin to realize what can be achieved using a liquid handling platform, such as the Freedom EVO, with 384and 1,536-well plates. It’s a whole new world,” concluded John. To find out more on Tecan’s Infinite microplate readers, visit www.tecan.com/detection To find out more about Chemical Biology at McMaster University, visit www.chemistry.mcmaster.ca/undergraduate-

“Using the Infinite microplate readers is a revelation for our students. After spending

Left to right: William Denk, Anya Todic, John Brennan and Kevin Yin using the Infinite M200 PRO microplate reader

Readers TECAN JOURNAL 1/2013

Investigating protein targets and cellular pathways in yeasts Scientists at the Donnelly Centre for Cellular and Biomolecular Research have chosen generations of Tecan microplate readers to monitor the effects of environmental or drug perturbation on molecularly barcoded yeasts.

The Donnelly Centre for Cellular and Biomolecular Research, based at the University of Toronto, Canada, is an interdisciplinary research institute that houses scientists from a wide variety of backgrounds, integrating the fields of biology, computer science, engineering and chemistry, as well as leading areas of biomedical research. Corey Nislow, Associate Professor at the Donnelly Centre’s Banting and Best Department of Medical Research (BBDMR) faculty, explained: “The Donnelly Centre houses faculty from many different departments, adopting a multidisciplinary approach to experimental studies with the intention of cross-pollinating between bioinformaticians, genomic and proteomic scientists. My own laboratory, and the laboratory of my collaborator Dr Guri Giaever, is responsible for running a next generation chemogenomics facility, and our work involves monitoring the growth of large pools of molecularly barcoded yeast, simultaneously screening 6,000 different mutants. The presence of a unique mutant barcode identifier enables us to distinguish each one of the mutants and deconvolute a complex sample pool at the end of an

experiment. This simplifies the procedure, allowing the pool to be treated as a simple culture and challenged with different environmental or drug perturbations.” Corey continued: “Guri, Michael Proctor (a research scientist working at Stanford University at the time) and myself first developed a workstation for this in 2000, consisting of four Tecan GENios™ microplate readers to monitor the growth of the culture, integrated with a PerkinElmer multiprobe. The basic principle is the same for this and the second generation system we later created using Tecan’s Safire™ reader and Freedom EVO® 200 liquid handling workstation. Based on a predetermined parameter, the liquid handler samples and reinoculates the culture, keeping it in logarithmic phase for up to 100 generations and allowing subtle effects on different mutants to be investigated. Initially, the liquid handler moves some of the culture to fresh media and, at the time of transfer, saves a sample so that the abundance of each strain can be decoded at the end of the experiment by microarray hybridization or next generation sequencing. The

“ We didn’t just ‘settle on’ Tecan, we purposely selected the Company.”

Corey Nislow and Guri Giaever with their Freedom EVO liquid handling workstation

Readers TECAN JOURNAL 1/2013

readers monitor both optical density and fluorescence – 95 % of the time we monitor optical density, but occasionally we study the readout from a fluorescent reporter – and each well of each plate is independently monitored. Essentially, we chart the abundance of every strain under a particular condition and, based on the abundance of the different mutants, infer the particular protein targets or cellular pathways that are important for culture survival under those circumstances. In the absence of these protein targets or cellular pathways, the culture is sensitive to that particular condition.” “For the first system we chose the GENios reader because it was the only instrument at the time that had a sufficient orbit to keep yeast cells well suspended and that could maintain temperature without condensation. However, although this system is still in use, the workflow means that the readers are effectively serving as shaking incubators, operating 24 hours a day, and shaking and reading at 15 minute intervals. The process of ejecting each plate to allow the liquid handler to sample the culture and return the plate takes a minute each time and is very much a rate-limiting step. In contrast, the second generation screening system is equipped with six shaking incubators, and the Safire microplate reader is just used to read! The speed of the Safire reader allows six to eight plates to be accommodated without any additional waiting time, and we can now interrogate model organisms that require light, such as the model algae Chlamydomonas; this would previously have been impossible in the dark of the reader. In addition, using dedicated shaking incubators has enabled us to increase capacity.” “We didn’t just ‘settle on’ Tecan, we purposely selected the Company, and so did all of our collaborators, who loved the growth curves they saw from our work. Every time we ran large screens we needed to confirm individual strains, and that required a server rack full of readers. We have remained with Tecan

and, between our Stanford site and Toronto, have 24 GENios systems, which are all still running! The flexibility of the Tecan systems is a big advantage, particularly the smooth information transfer.”

To find out more on Tecan’s detection solutions, visit www.tecan.com/detection

“As well as increasing throughput, our Tecan instruments have given us new avenues of exploration. We are now focusing on data collection and the introduction of new organisms, developing simple barcoding strategies for other organisms and investigating E. coli strains associated with Crohn’s Disease. We also plan to study more model organism genomes, de novo genomes, and will be doing a lot of next generation sequencing library preparation. In the future, we plan a third generation system, with two new generation Tecan readers and twelve shaking incubators, which will further increase our capabilities,” concluded Corey.

The GENios and Safire systems have been superceded by the Infinite® range of microplate readers, which features even more enhanced capabilities and is ideal for the assays and methods described above.

To find out more about the Donnelly Centre, visit tdccbr.med.utoronto.ca

Genomics TECAN JOURNAL 1/2013

Automated screening for recessive disorders elevates sensitivity to new heights Carrier screening based on next generation DNA sequencing has been completely automated at Good Start Genetics® Inc., with proprietary chemistry and customized protocols on six Freedom EVO® platforms giving significantly better turnaround times and increased reliability and reproducibility.

“ We chose our first Tecan system because it was one of the more customizable and readily adaptable systems available… we still feel the same today.”

Greg Porreca, Pat Saunders and Mark Umbarger with the Freedom EVO

Good Start Genetics, Inc., Cambridge, Massachusetts, is an innovative molecular diagnostics company that offers carrier screening for recessive genetic disorders, launching its pre-pregnancy screening service across the USA in 2011. Dr Mark Umbarger, Associate Director of Technology at Good Start, explained: “We use next generation DNA sequencing to assess whether patients referred to us by in vitro fertilization (IVF) clinics are carriers of certain recessive genetic disorders. The specific disorders tested are tailored to the patient, as directed by the physician. The panel of pre-pregnancy tests includes all 14 disorders recommended by the American Congress of Obstetricians and Gynecologists and the American College of Medical Genetics. Next generation sequencing (NGS) is allowing us to look deeper within each disease-associated gene and therefore enables us to identify a larger

number of disease-causing mutations. This comprehensive approach allows us to provide a more sensitive screen than is currently on the market. For example, we can detect ~500 cystic fibrosis disease-causing mutations while conventional genotyping-based approaches typically detect ~100 mutations.” Mark continued: “When it came to automation, we spoke to a number of other companies that were already using Tecan platforms and they recommended these systems.” Pat Saunders, Molecular Biology R&D Engineer at Good Start, added: “We chose our first Tecan system because it was one of the more customizable and readily adaptable systems available and, despite keeping a close eye on the market, we still feel the same today. We now have six Freedom EVO platforms – three Freedom EVO 200s and three

Freedom EVO 150s – purchased over the last two years. All the platforms have eight channel Liquid Handling Arms and PosID™ modules, and are adaptable to both 96- and 384-well microplate formats. Some are equipped with a Robotic Manipulator Arm, others with a MultiChannel Arm™ 96 for assay development.” “All of our assays are automated on the Tecan instruments, starting with DNA extraction from blood through to the genetic assays. There is virtually no hands-on pipetting, with full integration and automation of all steps, including library construction. The platforms in each of our laboratories – development, DNA extraction, pre-PCR and post-PCR – are configured accordingly.” “Sample preparation, such as that for library construction, molecular barcoding and NGS

TALK TO TECAN TECAN JOURNAL 1/2013

on our Illumina® HiSeq™ System are all done using proprietary chemistry developed in house, and fine-tuned, highly customized protocols,” Pat continued. “Some assays are completed in less than eight hours, others take a bit longer. All our tests go through validation on two levels – analytical, for accuracy, and clinical, to assess robustness in the production clinical laboratory. The validation process for our NGS assay on the Tecan platforms took multiple months and comprised simultaneous sequencing of hundreds of samples – target capture, molecular barcoding, sequencing and analysis – using an NGS-based approach, and also by an alternative reference method. We compared the results from hundreds of samples from both methods and found we had a very high specificity and sensitivity.” Mark concluded: “For Good Start Genetics, the three major benefits of the Tecan systems are reliable and walkaway automation, high throughput and reproducibility. We are constantly innovating or developing new approaches, improving current methods and coming up with new genetic tests, so the platforms’ flexibility is really important. We have been able to customize each system to our requirements and everyone in R&D and Clinical Operations at Good Start is very happy with them.” To find out more about Tecan’s genomic solutions, visit www.tecan.com/genomics To find out more about Good Start Genetics, visit www.goodstartgenetics.com

Nicholas Smith, Head of Product and Strategic Marketing Partnering Business

Leading the debate Tecan’s Partnering Business has a long history as a skilled and reliable original equipment manufacturer (OEM), bringing many years of expertise to the development of instrumentation for partnering companies. Our direct customers in the OEM business are usually diagnostic companies who approach Tecan with a specific need, looking for a partner to develop a solution for them. But what makes a company stand out as the OEM partner of choice? Clearly a system must be developed at the right cost, with the required functionality, and in the specified timeframe, but Tecan stands out from the crowd, offering more than just the basic necessities. Developing an understanding of our customers’ customer – the end-user of the instrument – is the key to that. By being proactive, we add even more value to OEM relationships by understanding our customers’ customers as well as they do, anticipating the future demands of the industry and using our expertise to suggest new approaches and technologies to make our customers more competitive in the market. Activities such as engaging with key opinion leaders, attending relevant conferences and establishing our own focus groups, for example the Tecan Symposium held recently in Boston on the subject of mass spectrometry (see pages 6-7), all contribute to our in-depth knowledge of the field. Our dedicated and experienced team continues to develop a better understanding of end-user applications, enabling Tecan to build superior solutions for its direct partners and ultimately benefitting their own customers – the end-users – as well. It’s a win-win situation. Email talk@tecan.com to tell us what you think differentiates Tecan from other original equipment manufacturers.

EVENTS 2013 TECAN JOURNAL 1/2013

Meet Tecan at these events Americas SLAS2013 – Society for Laboratory Automation & Screening

Orlando, FL, USA

12 – 16 Jan 2013

MSACL 2013 – The Association for Mass Spectrometry: Applications to the Clinical Laboratory

San Diego, CA, USA

10 – 12 Feb 2013

MD&M West – Medical Design & Manufacturing

Anaheim, CA, USA

12 – 14 Feb 2013

Molecular Med TRI-CON 2013

San Francisco, CA, USA

13 – 14 Feb 2013

AAFS 65th Anniversary Meeting – American Academy of Forensic Sciences

Washington, DC, USA

18 – 23 Feb 2013

SOT Annual Meeting and ToxExpo™

San Antonio, TX, USA

10 – 14 Mar 2013

Pittcon™ Conference & Expo 2013

Philadelphia, PA, USA

18 – 21 Mar 2013

245 ACS National Meeting & Exposition

New Orleans, LA, USA

07 – 09 Apr 2013

Ninth Annual PEGS – the essential protein engineering summit

Boston, MA, USA

29 Apr – 03 May 2013

The 2013 Annual Meeting of Japan Society for Bioscience, Biotechnology & Agrochemistry

Sendai, Japan

24 – 27 Mar 2013

CMEF China International Medical Equipment Fair Spring 2013

Shenzhen, China

17 – 20 Apr 2013

ISBER 2013 Annual Meeting & Exhibits

Sydney, Australia

05 – 09 May 2013

Functional Genomics and Proteomics – Applications, Molecular Diagnostics & Personalized Medicine

Frankfurt, Germany

31 Jan – 01 Feb 2013

5th Congress of the Spanish Proteomics Society

Barcelona, Spain

05 – 08 Feb 2013

3 Annual Cell Culture World Congress

Munich, Germany

26 – 28 Feb 2013

Advances in Microarray Technology

Barcelona, Spain

05 – 06 Mar 2013

ARABLAB 2013

Dubai, UAE

10 – 13 Mar 2013

World Immune Regulation Meeting-VII

Davos, Switzerland

13 – 16 Mar 2013

LabAutomation

Netherlands

09 Apr 2013

Asia and Pacific

Europe, Middle East and Africa

Tecan are pioneers in automated liquid handling and innovative life science solutions. For over 30 years we continue to enable and support our customers to make the world a healthier and safer place.

Headquarters: Tecan Group Ltd., Seestrasse 103, CH-8708 Männedorf, Switzerland T +41 44 922 88 88 F +41 44 922 88 89 info@tecan.com

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Tecan Group Ltd. makes every effort to include accurate and up-to-date information within this publication, however, it is possible that omissions or errors might have occurred. Tecan Group Ltd. cannot, therefore, make any representations or warranties, expressed or implied, as to the accuracy or completeness of the information provided in this publication. Changes in this publication can be made at any time without notice. All mentioned trademarks are protected by law. In general, the trademarks and designs referenced herein are trademarks, or registered trademarks, of Tecan Group Ltd., Mannedorf, Switzerland. A complete list may be found at www.tecan.com/trademarks. Product names and company names that are not contained in the list but are noted herein may be the trademarks of their respective owners. For technical details and detailed procedures of the specifications provided in this document please contact your Tecan representative. This journal may contain reference to applications and products which are not available in all markets. Please check with your local sales representative: www.tecan.com/contact