Tecan Journal Edition 02/2005 by Tecan Group Ltd.

ISSN 1660-5276

Tecan Journal Autumn/Winter - Edition 2 2005

25 years in the making: a brief history of Tecan page 2

High throughput scrapie genotyping page 10

Solving protein structures on a high throughput scale page 20 Cover shows Dr Mark Wang, Chief Representative, Tecan Group Ltd., Beijing Representative Office and Patrik B端chli, IT Manager Operation, Tecan Group Ltd., Switzerland.

25 YEARS TECAN

25 years in the making: a brief history of Tecan Year, and awarded the prestigious BrancoWeiss Prize. Newspaper articles of the day paid tribute to Tecan’s distinctive spirit and unique values of open communication with staff, absolute dedication to the task at hand, willingness to innovate, flat hierarchy and high degree of job satisfaction achieved through individual responsibility. Tecan Holding was founded in 1986, at the same time as Tecan Asia (PTE) opened in Singapore. By then, Tecan employed a workforce of 80, generated sales of almost CHF 20 million, boasted an exceptional product range and was spread across ten buildings in Hombrechtikon.

Expansion: from private company to the stock exchange

The old Tecan building in Hombrechtikon

Pioneer days: from a Swiss mountain farm to the world stage Tecan was founded in 1980 by four engineers in the Swiss village of Hombrechtikon, with the aim to develop measurement, analysis and laboratory automation devices. The first offices were located in a renovated farmhouse, the family home of CEO and founder, Heinz Abplanalp. He and his founding colleagues, Heini Maurer, Heini Möckli and Gallus Blatter, were soon joined by two further partners, Daniel Ryhner and Martin Stoffel. In the beginning, the focus was exclusively on development, but the company’s success soon led Tecan to begin manufacturing proprietary instruments and OEM products for laboratory

Tecan Journal 2/2005

automation and environmental analysis. Early production took place in renovated barns, houses and warehouses scattered around the village. Soon afterwards, in 1982, Tecan opened a sales and marketing unit for North America, Tecan US, based at Chapel Hill, North Carolina, and Tecan Production Corp., a manufacturing unit for the US market. When the company faced liquidity problems in 1984, the financing company, Zelux AG, bought a 48% share in the upand-coming enterprise. Two years later, Tecan was voted Swiss Company of the

In 1987, Tecan floated on the Zurich stock exchange and all employees received a stake in the company through a distribution of shares. The share price skyrocketed and, thus equipped with sufficient resources, Tecan subsequently purchased ten companies, including the US-based OEM manufacturer Cavro Scientific Instruments Inc. The following year, 1988, Tecan acquired SLT Laborinstrumente GmbH, based in Salzburg, Austria. By the end of 1988, Tecan Holding already comprised 25 companies. It soon became clear, however, that growth and costs had spiraled out of control. By mid 1989, the Tecan Group was on the brink of bankruptcy and trading in their shares had to be temporarily suspended. The renowned crisis manager and former manager at Bührle, Hans Widmer, stepped into the breach, joining the company

25 YEARS TECAN

Tecan voted as Swiss Company of the year and awarded the Branco-Weiss Prize; TV broadcast from Nov 24 1986

Tecan AG Annual Report 1995 The 25 years’ celebration at Tecan Austria

as CEO. He was to introduce capital strengthening measures and restructure the company. Tecan’s management resolved to sell almost all of the 25 companies in the Tecan Group, retaining only those profitable firms involved in automated liquid handling (Tecan AG, Cavro and SLT) as well as sales operations in Germany, the UK and the USA. Restructuring was rapid and painful, but it was the right medicine, and the Group recovered quickly. Thanks to the unrelenting dedication of its staff, the Tecan Group was making a profit again by 1990. The founder, Heinz Abplanalp, then left the company, and Peter Bollmann took over as CEO.

Consolidation: shoring up success The remaining founders also left the company in 1991. The introduction of Charter 91 marked a renewed emphasis on Tecan’s corporate culture and this document put forward that employee satisfaction was just as important a criterion for the company as financial success and customer service. At the same time, Charter 91 demanded transparent procedures and effective controls.

being introduced and costs were being optimized. Moreover, Tecan introduced more efficient information and project management systems. In 1995, investment in R&D swelled to 14% of revenue, and the Genesis® series was launched. Tecan received a further boost in 1998 when a sales promotion agreement was signed between the Tecan Group and Abbott Laboratories for the marketing and sale of the Genesis FE 500 Workcell.

When Gottlieb Knoch became CEO, President of the Board and majority shareholder in 1992, the Tecan Group already employed 330 people and was returning ever improving results. The new sales unit in Japan was also proving profitable. The quality management processes had acquired ISO 9001 certification at all three manufacturing companies within the Tecan Group by 1994. A proportion of revenue (10.4%) was being invested in R&D, innovative production methods were

Mr Widmer in front of the old Tecan building

Tecan Journal 2/2005

2G5L OY B EAR L SNTEEWCSA N

Tecan at the AACC Annual Event in USA (Orlando, Florida)

In 1997, the American PerkinElmer Corporation purchased a majority of voting shares from Gottlieb Knoch. PerkinElmer’s strength in chemicals and instruments for pharmaceutical and biotech laboratories gave the Tecan Group a leg up into the Life Sciences market. PerkinElmer’s sale of its shares on the open market in 1999 led to a restructuring of the company, followed in June of the same year by an IPO and listing on the Zurich stock exchange. The IPO was overseen by a new CEO, Emile Sutcliffe. The company’s top management subsequently launched the Tecan 2000 program, and adjusted the Group’s strategic orientation.

In 2003, CEO Emile Sutcliffe left the company. After an interim phase, during which Board Chairman Mike Baronian headed operations, Aitor Galdos took over at the helm late in the summer of 2003. He immediately launched a comprehensive program to evaluate the corporate strategy which was then complemented by a second comprehensive program for the assessment of all regulatory issues. A number of adjustments are being made within Tecan’s organization as a result of this gap analysis. At the end of 2004, the then CEO Galdos left the company due to irreconcilable differences with the Board of Directors.

Turbulent times: from the stock market boom to a sobering wake-up On the back of the re-orientation and a generally positive market environment, Tecan’s share price initially steamed upwards from 1999. But, when the economy hit a downturn not long thereafter and the stock market tumbled, it became clear that the rapid change initiated by the company had not been without its ill effects. Problems in production and distribution logistics, aggravated by a patchy management information system and bad investments in certain new technologies, caused Tecan to steadily lose ground to its competitors. The team from Tecan Austria Tecan Journal 2/2005

The future: strength and growth through efficiency, continuity and acquisitions The new management chief, Thomas Bachmann, who was appointed in February 2005, has prescribed a new program for the company in recent months. The first stage of this program shall aim to stabilize declining revenues and profitability. Once this has been achieved, the strategy will focus on securing a long-term and sustainable turnaround. The new program is centered around the following activities:

25 YEARS TECAN

1985 – Extract of Tecan Product range: pH-Electrodes, Tecan Plant SAP Extractor 400, Tecan RSP 5052, Tecan Samplers 500 Series

• improved customer focus throughout the company and better service standards • systematic implementation of the quality improvement measures that have been introduced with regard to regulatory matters • simplification of the organizational structures and strengthening the management • a more professional approach to internal visibility and reporting • regaining trust by means of reliable communication with staff, customers, shareholders and the financial market • focusing project and service innovations on promising projects in the four areas: components business, detection devices, automated liquid handling and customer services • improved cost awareness and management This is designed to fuel an increase in the company’s profitability and lead off a new growth phase. As well as aiming to generate organic growth (at least in line with market growth), Tecan’s Board of Directors and management are also planning to boost external growth opportunities by means of company acquisitions. One of the first of these was the purchase of REMP in June 2005.

REMP is located in Oberdiessbach, near Berne. The company was founded in 1986, employs a workforce of 154 and generated sales of CHF 36.4 million in 2004. REMP is the world leader in the manufacture of large-scale automated storage and retrieval systems for biopharmaceutical companies. REMP also produces devices for sealing, decapping, identification and sorting of samples, as well as consumables for fast, reliable sample logistics. The devices produced by both REMP and Tecan cover a large part of the drug discovery process and are also suitable for use in new applications in the areas of forensics and clinical diagnostics in the future. In July 2005, Tecan’s management decided to abandon the LabCD program and close its site in Boston. These moves finally close the chapter on the meager success of the development project that Tecan inherited when it bought over Gamera Bioscience.

Despite the recent years of turbulence, Tecan still boasts a sound financial base and generates a high cash flow which is intended to facilitate further strategic steps in the future.

Conclusion: leaving rough seas behind, with new land in sight Since its formation in 1980, Tecan has weathered many storms, but one thing has remained constant throughout all of these difficult periods: the Board of Directors, the management and the staff have pulled together and overcome all obstacles. With the growth recorded in recent months, the Tecan team has demonstrated that it will, once again, uphold the tradition of success left by earlier Tecan generations.

L AT E ST P R O D U C T S

Latest product bulletin more instrumentation to improve our customers’ laboratory automation

Two LiHa arms with fixed or disposable tips

The Freedom EVO with dual liquid handling arms The two LiHa arms can work in series or independently

Freedom EVO® Freedom EVO® automated workstations are now available with a second liquid handling arm (LiHa) to improve both the speed and flexibility of the platform and enhance the liquid handling power of any laboratory.

InfiniteTM 200 The new Infinite™ 200 is Tecan’s groundbreaking series of flexible, upgradeable multi-mode microplate readers, offering a choice of monochromator or filter system and configurable detection modes as well as optional injectors to allow users to tailor the instrument for their individual needs. The Infinite 200 upgradeable multi-mode microplate reader

Tecan Journal 2/2005

CO N T E N T S

Te-PSTM The Te-PS™ is based on Tecan’s standard high precision LiHa and is equipped with a laser guided positioning sensor that guarantees reliable and accurate routine use of high density pipetting formats, such as 1536-well plates.

Special 25 years in the making: a brief history of Tecan Special feature detailing Tecan’s impact on the life science industry over the last 25 years page 2 Latest product bulletin and contents at a glance page 6

Measuring calcium-triggered luminescence in CHO mito-Photina™/A3 cells Using the GENios™ Pro microplate reader to measure flash luminescence in a reporter gene assay page 14 Two heads are better than one! Tecan and REMP complement each other perfectly in the new partnership that will benefit all life science laboratories page 16

Freedom EVOlyzer® – successful ELISA automation at Swiss Reference Lab Tecan solves the ELISA throughput bottlenecks for a large clinical laboratory page 8

Freedom EVO® – complete automation of genotyping assays at Kiel The Freedom EVO 384R package increases the throughput of an automated genotyping laboratory page 18

Te-PS (Tecan Positioning System) uses a laser based positioning system to allow robust and reliable tip positioning in high density formats.

Te-PoolSafeTM Coming soon: Tecan’s new Pool Safe Option, Te-PoolSafe™, is a balance module that measures and checks every liquid arrival to each sample in a set of tubes, bringing enhanced safety and security to all pooling and testing applications. Find out more on page 22

High throughput scrapie genotyping An automated method for large volume identification of scrapie-susceptible sheep page 10

Identifying genetic causes of learning disability Array-based comparative genomic hybridization on the HS 4800TM Hybridization Station reveals previously unidentified chromosomal imbalances in patients with learning disability page 12

Solving protein structures on a high throughput scale Developing automated applications for all aspects of protein characterization, including a dot blot, on Tecan’s liquid handling platforms page 20

New products and customer support Discover more about our new products, including disposable reagent troughs, dual liquid handling arms, the InfiniteTM 200 readers and more innovations to make your life easier page 22

Tecan Journal 2/2005

A P P L I C AT I O N D I A G N O ST I C S

Tecan’s Freedom EVOlyzer

successful ELISA automation at Swiss Reference Lab As the largest hospital-based clinical laboratory in north-east Switzerland, the Institute for Clinical Chemistry and Hematology (IKCH) in St Gallen serves an area of 450,000 inhabitants in all fields of laboratory medicine, including clinical chemistry, hematology, coagulation, immunology and molecular genetics. The Institute’s clients include public and private hospitals, general practitioners and private laboratories, as well as the pharmaceutical industry. Although the laboratory already used a number of automated analyzers, these systems did not cover the complete range of the laboratory’s assays and therefore the IKCH had been looking for ways to expand their automation.

The loading interface uses integrated sensors and LED lights to guide and monitor user actions.

Sample and reagent loading is guided and monitored by the loading interface

A Freedom EVOlyzer 150 with 8 standard tips, the configuration used at IKCH in St Gallen

The team at IKCH looked at several ELISA automation platforms available on the market, but found that most were one or two needle systems, not suitable for the majority of their assays. Professor Walter Riesen, Head of IKCH, explained that they had a difficult time to find an instrument that could meet their special needs: “Firstly, it had to be an open platform that could manage our constantly growing and changing panel of tests. It also had to be able to simultaneously process multiple assays and, very importantly, it had to be able to handle assays with short room temperature incubation times." Tecan Journal 2/2005

This last requirement proved a particular problem for many of the automated ELISA systems that the IKCH tested. Most one or two needle instruments were unable to handle short room temperature incubation times required for some target assays, which meant having to divide 96-well plates, actually increasing rather than decreasing handson time for those assays. “Simultaneous performance of multiple assays with short incubation times was a big problem,” explained Dr Wolfgang Korte, senior staff member and project leader at IKCH. “But, when we found that Tecan’s Freedom EVOlyzer® 150 could be equipped with up to eight needles, we gave it a try. We tested the system on a Chromogenix “COALIZA® Protein S – Free” assay, which involves some tricky predilution of the calibrators and a ten minute room temperature incubation step at the end of the process. We realised that the Freedom EVOlyzer could handle the assay to our full satisfaction.”

Freedom EVOlyzer – a new generation of ELISA analyzers The Freedom EVOlyzer is Tecan's new generation of IVD-D (98/79/EC) compliant ELISA analyzers that replaces and builds on 15 years’ experience of the successful RMPTM and MinilyseTM/MiniSwiftTM systems. Like its predecessors, the instrument is available in several worktable sizes (100, 150 and 200 cm). The instrument can be equipped with two, four or eight needles, using either steel or disposable tips. The liquid handling includes single and multipipetting, parallel sample pipetting over several plates, complete flexibility in predilution and high safety using liquid level and clot detection. The independent robotic manipulator (RoMa) arm for plate transport allows simultaneous use of the system’s different devices, including up to 12 chambers for heated incubation as well as 12 chambers for room temperature incubation. The Freedom EVOlyzer relies on the well known, reliable Columbus™ plate washer and Sunrise™ absorption reader, and the integrated Positive Identification (PosID™) barcode reader identifies samples, plates and reagents.

A P P L I C AT I O N D I A G N O ST I C S

The highest possible levels of throughput, safety and flexibility have been target objectives during the development of the system. Factors including continuous loading, parallel sample pipetting on up to six plates, as well as configurable worktable layouts all add to its appeal. Enhanced safety features include extensive use of safety panels, door locks, sensors, lights and acoustics to enable close monitoring of all user and instrument actions. The wizard-driven, touch screen compatible easy-to-use Run Control Software guides the user through the daily operation. Powerful editing programs extend the flexibility even further, allowing skilled users to leverage maximal performance from the new software.

“One of our main objectives was to reduce our hands-on time and running multiple assays in parallel leaves our employees free for more qualified tasks. We needed an instrument that is easy to operate, that provides us with clear instructions and that, once started, we can confidently leave alone to get on with the assays unattended. After all, reliability and walk-away capability is the key to automation. The Freedom EVOlyzer should allow us to reach these goals. The system’s ability to run multiple assays in parallel clearly allows us to shorten our turnaround time on our assays.” IKCH and Tecan have now embarked on a close collaboration that will continue the development of the Freedom EVOlyzer system. Dr Korte concluded,

“

Dr Korte and Mrs Aichele at the IKCH with their Freedom EVOlyzer

Chromogenix – COALIZA® Protein S - Free assay Summary of procedure Serial predilution of Calibration Plasma Cal1 1.0

IU/mL free PS

10 µl of Cal Plasma

1000 µl of diluent

Cal2 0.5

IU/mL free PS

500 µl of Cal1

500 µl of diluent

Cal3 0.25

IU/mL free PS

500 µl of Cal2

500 µl of diluent

Cal4 0.125 IU/mL free PS

500 µl of Cal3

500 µl of diluent

Predilution of samples 1:101 Add diluted sample, control and calibrators 100 µl/well Add conjugate 100 µl/well Incubate: 45 min at RT Wash 4 x 300 µl Load freshly prepared substrate-chromogen TMP dilution Add substrate 200 µl/well Incubate: 10 min at RT Add Stopping Solution 100 µl/well Read at 450 nm

We are confident that we have made the right choice and we look forward to the future collaboration with Tecan.

COALIZA®

And the overall flexibility that the Freedom EVOlyzer offered was a major priority, as Dr Wolfgang Korte explained: “We were not particularly aiming to run high throughput assays on the instrument, but instead, will be running a large number of tests in parallel with only a moderate number of samples. We very much wanted to automate as many of our applications as possible to a microtiter plate format and needed a system that is flexible enough to handle these different assays.”

COALIZA® is a registered trade mark of Chromogenix. Freedom EVOlyzer® is a registered trademark of Tecan Group Ltd., Männedorf, Switzerland

”

Tecan Journal 2/2005

A P P L I C AT I O N B I O P H A R M A

High throughput scrapie genotyping: a best practice automation example with 8,000 samples processed per day Stefan Sidler, Laboratorio Central de Veterinaria, Subdireccion General de Sanidad Animal, Ctra. Algete Km 8, 28110 Algete (Madrid), Spain

Primary sample distribution with a Tecan Genesis Freedom 150/8 pipetting platform

Scrapie is a fatal, neurodegenerative prion protein disease of sheep and goats that belongs to the family of transmissible spongiform encephalopathies (TSEs). European Union legislation, introduced in April 2005, requires that every member state establish a program to breed out scrapie-susceptible sheep. This presents an interesting challenge that can be solved using scrapie genotyping to select animals for these breeding programs. A number of single nucleotide polymorphisms (SNPs) have been discovered within PRNP, the prion protein-encoding gene, that affect the susceptibility of an individual sheep to scrapie. An animal’s risk, following contact with the scrapie pathogen, can be classified based on a single nucleotide extension (SNE) assay that determines four specific SNPs in the PRNP gene.

Identifying animals with high susceptibility to scrapie The Laboratorio Central de Veterinaria is the Spanish reference lab for scrapie genotyping and reports directly to the Ministry for Agriculture, Fishing and Feeding (Ministerio De Agricultura, Pesca y Alimentación; MAPA). Here, we have developed a high throughput, automated procedure for genotyping in 384-well plates that enables us to process about 8,000 samples per day and 1.8 million samples per year.

Table 1 Tecan Journal 2/2005

Our lab receives barcoded primary blood samples in rubber stoppered plastic tubes with a minimum volume of 2.5 ml blood. All data relating to animals, owners, flocks etc. are stored in a central database (ARIES) by the field veterinarians and these data are downloaded into our LIMS when the samples arrive. The blood genotyping protocol consists of eight steps and the whole workflow takes about 10 hours in total (Table 1). Primary sample distribution Two Genesis® Freedom 150/8 pipetting platforms are used for the primary sample distribution, each equipped with an 8-tip LiHa arm, a washing station with fixed tips and a PosIDTM System for automated barcode reading (Fig 1). The pipetting platforms process 384 blood samples per run and the LiHa arms are equipped with fixed steel tips which pierce the rubber stoppers, preventing unnecessary opening

A P P L ICCU ATSTO ION M EBRI OSPUHPAPROMRT A

Figure 2: DNA extraction and PCR setup with a Tecan Genesis Freedom 200 pipettor with centrifuge (left, under the worktable; long RoMa arm above), Te-MO 3/5 multi-pipettor with a 96-channel pipetting head (middle) and storage hotels (right)

Figure 3: PCR cleanup and setup of SNE reaction with a standalone Tecan multi-pipettor Te-MO 3/5 with a 384-channel pipetting head and an integrated vacuum manifold

of the primary tubes before sample distribution. Samples are identified by the PosID and distributed into tubes in 96-well racks for storage and into 384 half deep well plates (20 µl of each blood sample) for the subsequent DNA extraction.

to be dispensed without contact with the samples, to prevent contamination or carryover. Disposable tips, DNA binding plates, PCR plates and other plasticware used during the protocol are placed in plate hotels and transported to the Te-MO 96 by the short RoMa arm.

DNA extraction, PCR setup of the SNE reaction The DNA extraction is performed on two Genesis Freedom 200 platforms, each equipped with a long and a short RoMa arm, Tecan’s Multichannel Pipettor Te-MO™ 3/5 with a 96-channel pipetting head (Te-MO 96), and a centrifuge below the worktable which can be automatically loaded using the long RoMa arm (Fig 2). Both standard steel tips (96-tip block) and disposable tips are used during the DNA extraction as the Te-MO 96 can exchange disposable and fixed tips within one run. The 96-tip block is used during DNA washing steps, where reagents need

Figure 4: Typical scrapie genotyping result visualized with the GeneMapper software (Applied Biosystems). Each peak corresponds to one of the SNE products generated from the four SNP sites which have been studied. In this example, all SNP sites are present in homozygous form since there is only one peak per SNP site

The Tecan Genesis Freedom 200 pipetting platforms are also used to perform PCR setup, the 384-well PCR plates are then manually sealed and PCR is performed with standard 384-well thermocyclers. PCR cleanup, SNE assay setup and shrimp alkaline phosphatase enzymatic purification of the SNE products are carried out on two Te-MO 3/5 multipipettors with 384-channel pipetting heads (Te-MO 384) and an integrated vacuum manifold (Fig 3), with standard 384-well thermocyclers used for the reaction steps. Laser-induced fluorescence capillary electrophoresis (LIF-CE), revision of genotyping results and export of results to ARIES Products generated from the SNP sites are detected by LIF-CE on two 96-capillary 3730xl DNA Analyzers from Applied Biosystems, followed by automated allele-calling using GeneMapper software (Applied Biosystems) (Fig. 4) . Each genotyping result is visually controlled and approved by experts prior to being exported to ARIES.

Conclusion We currently have two parallel working Tecan platforms performing high throughput genotyping, allowing us to process over a million and half samples per year and, to our knowledge, this means our lab has the highest automation level in the EU for this application. In Spain, a large number of breeder organizations have already introduced genetic selection into their breeding programs based on scrapie genotypes. It is important that those breeders can identify their animals’ genotypes in the shortest period of time to optimize breeding management and, with the high throughput automated procedure described here, our lab can provide more than 95% of the results within two weeks. We chose the Tecan instruments because they offer the reliability, durability and flexibility necessary for this application, and they are extremely compatible with the 384-well format, which also allows us to save costs on consumables. Our high throughput genotyping equipment also have the potential for use in a variety of other genotyping applications. Genesis® Freedom, Freedom EVO® are registered trademarks of Tecan Group Ltd., Männedorf, Switzerland Applied Biosystems® is registered trademark of Applera Corporation or its subsidaries in the U.S. and/or certain other countries. GeneMapper™ is trademark of Applera Corporation or its subsidaries in the U.S. and/or certain other countries.

Tecan Journal 2/2005

AU TO M AT E D L I Q U I D H A N D L I N G

Identifying genetic causes of learning disability Learning disability affects 1 to 3% of the population and, although the underlying origins remain unidentified in 30 to 50% of cases, constitutional chromosomal imbalance is the single most common known cause, found in 4 to 28% of patients. However, classical techniques for cytogenetic studies, such as karyotype analysis, have only limited resolution and are not sufficiently sensitive to detect subtle chromosomal rearrangements involving fewer than 5 Mb. Dr Nigel Carter’s team of molecular cytogenetics researchers at The Wellcome Trust Sanger Institute in Hinxton, Cambridge (UK), has developed high resolution microarrays for genome-wide screening of chromosomal imbalances in patient DNA samples by comparative genomic hybridization (CGH). The information provided by array-based CGH (aCGH) can be directly linked to the existing sequence of the human genome. From these studies they hope to identify previously undetected microdeletions and microduplications that are responsible for learning disability and dysmorphology. The investigations require analysis of large numbers of slides and, as carrying out these procedures manually can be highly laborious, the team looked into automating the process, as Dr Richard Redon – Research Associate in Dr Carter's laboratory – explained, “Hybridizing 24 slides manually takes one person a whole week whereas, with an automated system, this person can potentially complete 24 slides in just a day. Other advantages of automation include the fact that a robotic system consistently gives highly reproducible results and it is not prone to human error – a particularly important feature for clinical work.”

Tecan Journal 2005

The researchers chose a Tecan HS 4800TM Hybridization Station to screen samples from patients with learning disability and dysmorphology for deletions and duplications that may contribute to the phenotype, followed by samples from the patients’ families. “The main findings from this project were some previously undetected imbalances in 25% of the patients we analyzed, but there are two problems with these data. The first is that some of these imbalances are inherited, so we do not know how to interpret these deletions or duplications at the moment. The second problem is that we did not find any identical deletion or duplication, so each identified imbalance affected a different part of the genome,” said Dr Redon. The team has now embarked on a separate study to address the problem of inherited genetic imbalances, as Dr Carter described, “It is very important to know whether identified copy number changes are inherited from a normal parent because, if so, then it is less likely that they are the cause of the learning disability, even though that microdeletion or duplication may still be involved. We are also trying

to identify the natural variation in the normal population, because it has recently emerged that a large number of copy number variations exist within the genome of normal individuals.” The second problem with the data remains unexplained, but Dr Carter thinks that this is due to the ascertainment bias in the patients under study, “There are some more frequent phenotypes which clinicians can identify as being common across individuals with learning disability and dysmorphology and, when we look at these patients, we invariably find the same region has been involved in a deletion or a duplication. The patients in our present study are selected because they do not fit with these known syndromes and so we are looking at individuals with rare or unique changes in their genome, generating a rare profile of learning disability and other dysmorphic features. We now need to look at a larger number of samples to find patients in this group with similar affected regions, in order to make some inference about which genes are likely to cause the disease.”

AU TO M AT E D L I Q U I D H A N D L I N G

“We have also set up an international collaborative database called DECIPHER that enables laboratories around the world, doing similar sorts of aCGH studies, to submit their patients’ observed phenotype and genomic changes into a common database. This should help the identification of patients with similar copy number changes in their genome so we can then look at the genotype-phenotype correlation and, ideally, pinpoint the genes most likely responsible for the particular disease.” Although this database is very new, along with collaborators at the Hospital NeckerEnfants Malades in Paris, Dr Redon has already identified two individuals with very similar phenotype and deletions on chromosome 9, suggesting the existence of a previously unidentified syndrome. This is not only of interest to molecular cytogeneticists: defining specific syndromes allows clinicians to have a good idea of disease causes when they see patients with a similar phenotype. Following up these patients means clinicians are better able to counsel new families as to the likely outcome and long term prospects for the child.

“An affected family usually wants to know why a child has a particular problem and it is very helpful for the family to actually understand why the disorder has manifested. Furthermore, the particular chromosomal rearrangement can be screened for, allowing prenatal diagnosis.” Dr Carter and his colleagues now have three HS 4800 to cope with their workload,

“I think all three of the machines are used virtually non-stop throughout the week!” The applications described here are not available in the US outside of the research market. The above text does not imply endorsement of The Wellcome Trust Sanger Institute.

Understanding the genetic causes of learning disability and dysmorphology can benefit patients and their families in other ways, as Dr Carter explained: HS 4800™ is a trademark of Tecan Group Ltd., Männedorf, Switzerland

HS 4800 increases throughput and reproducibility in genome-wide screening of chromosomal imbalances by array-based CGH (aCGH)

Tecan Journal 2/2005

DETECTION

Measuring calcium-triggered luminescence in CHO mito-Photina™/A3 cells Mateja Niederreiter and Manfred Lansing Reporter gene assays have become an indispensable part of today’s life science research as measuring the activity of a reporter gene, which reports the activity of a gene of interest, provides a very informative and appropriate experimental tool for cell-based experiments. Such assays are especially useful for investigations into gene expression or signal transduction pathways within living cells. It is not surprising, therefore, that the use of reporter gene assays has increased and a considerable number of reporter gene assays have been developed in recent years. Here we describe flash luminescence measurements performed on Tecan’s GENios™ Pro, a multifunctional injector plate reader, with a receptor-ligand assay based on the Chinese Hamster Ovary (CHO) mito-Photina™ reporter gene technology (Axxam1). The CHO mito-Photina reporter gene technology has been designed for functional studies of molecular reactions

Tecan GENios Pro – highperformance injector reader

that are able to induce variations in the intracellular calcium concentration. Photina, the reporter gene, was successfully cloned into CHO cells to encode a recombinant, calcium-sensitive photoprotein. These photoproteins (e.g. aequorin) have calcium-binding sites and, upon binding of the calcium ions, the photoproteins immediately bioluminesce. In a reporter gene assay, this reaction can be applied to analyze various cellular processes associated with Ca2+-mediated signal transduction.

Figure 1: Measurement settings for luminescence measurements with CHO mito-Photina/A3 cells. All measurements were performed in the luminescence well kinetic mode

Tecan Journal 2/2005

Materials and methods Materials: CHO mito-Photina/A3 cells (Axxam), IB-MECA and ATP (Sigma), coelenterazine (PJK), Tyrode buffer (130 mM NaCl, 5 mM KCl, 2 mM CaCl2, 1 mM MgCl2, 5 mM NaHCO3 and 20 mM HEPES, pH 7.4), Triton buffer (25 mM TRIS, 25 mM Na2HPO4, 2 mM DTT, 10% (v/v) glycerol and 2% (v/v) Triton X-100, pH 7.8) and 384-well microtiter plates, white (Greiner). Assay protocol: dose response measurements were performed with IBMECA and ATP using 384-well microtiter plates as follows: mito-Photina/A3 cells were cultivated according to the cell culture protocol provided by Axxam1. Cells were seeded (~500 cells/well) and, after 24 h, the cell medium was replaced with 30 µl/well Tyrode buffer containing 5 mM coelenterazine. The plate was then incubated for 3 h at 37 °C prior to measurements, which were started by adding 30 µl/well 2x IB-MECA or 2x ATP solution, prepared in Tyrode buffer and always injected into the wells directly by the instrument. Cell samples treated with ATP were finally lyzed by injecting Triton buffer (30 µl/well). Measurement settings: luminescence (well kinetic), integration time: 1000 ms, number of cycles: 70 (IB-MECA) and 50

DETECTION

(ATP); well kinetic interval: minimum, injector A volume: 30 µl, injector A delay: injection before cycle 10, injector A speed: 100 µl/s, injection mode: standard.

Results and discussion

Figure 2: Dose response curves for IB-MECA

Figure 3: Dose response curves for ATP

Figure 4: Cell lysis after ATP addition

The CHO mito-Photina/A3 cells stably express the G protein-coupled cell membrane receptor protein Adenosine3 (A3) together with a chimeric Gα protein to switch the signal to the phospholipase C (PLC) pathway. Specific binding of a ligand to the receptor causes a cascade of G protein-triggered signaling events in the cell, including release of calcium from intracellular calcium stores. Our experiments were conducted by using (i) IB-MECA, a selective A3 agonist and (ii) ATP, which causes Ca2+ release on binding to the endogenously expressed P2Y2 receptor. Different ligand concentrations were applied to vary the signal intensity and the luminescence measurements were initiated by injection of 30 µl ATP or IB-MECA, respectively (Fig 1) and performed in the luminescence well kinetic mode. Depending on the ligand, the number of cycles was set to 70 (IB-MECA) or 50 (ATP) and each signal was integrated over the time period of 1 s. Dose response curves were used to correlate the signal intensity with the ligand concentration. As expected, the detected signal was intensified as the concentration of the respective ligand increased (Figs 2 and 3). An IB-MECA concentration as low as 1 nM was sufficient to trigger a detectable amount of light, whereas a corresponding ATP concentration lay within the range of 1 µM. All curves were detected in several replicates whereby reproducible results with high data quality were achieved. In addition to the dose response experiments, we measured the amount of light produced by mito-Photina when completely saturated with calcium ions. For that purpose, the cells were treated with ATP and the responding luminescence

was measured. After this first signal had declined, the cells were subjected to lysis with Triton buffer, which was injected directly into the wells. Again, flash luminescence was detected, but this time the signal was caused by binding of extracellular calcium to Photina, i.e. calcium that was included in the cell medium. The final analysis showed that the signal intensity after cell lysis strongly depended on the amount of ATP previously added to the cells and, therefore, on the grade of mito-Photina saturation with intracellular calcium. The brightest signal was obtained for cells without added ATP, whereas cells previously treated with ATP produced signals with intensities that were inversely proportional to the amount of added ATP (Fig 4).

Conclusion Using the GENios Pro for highly sensitive flash luminescence measurements, based on the mito-Photina reporter gene technology, resulted in extremely reproducible luminescence signals. These findings demonstrate the outstanding performance and suitability of GENios Pro for this type of measurement as well as the extremely robust reporting capabilities of mito-Photina based applications.

Reference 1. www.axxam.com

Acknowledgment We cordially thank Sabrina Corazza and Germano Carganico at Axxam for the excellent cooperation and valuable contributions to this work. CHO mito-Photina™ is a trademark of Axxam srl, Milan, Italy GENios™ Pro is a trademark of Tecan Group Ltd., Männedorf, Switzerland

Tecan Journal 2/2005

SAMPLE MANAGEMENT

Two heads are better than one!

The new agreement between Tecan and REMP is good news indeed for many of our customers. The product portfolios of both companies complement each other and, together, they comprehensively cover applications in the drug discovery, biotech, diagnostic and general laboratory arenas. REMP’s revolutionary large-scale automated storage and retrieval systems integrate seamlessly with Tecan’s state-of-the-art automation platforms and the affiliation between the two companies makes perfect sense.

Small-Size Store (SSS) with integrated Tube Punching Module (TPM)

REMP set new standards of quality for sample storage with the development of its REMP Tube Technology™ in a 384-well format. The introduction of individually sealed tubes and the concept of storing small aliquots ready for single use has eliminated unnecessary freeze/thaw cycles, carryover and dilution effects, as well as reducing the water take-up and exposure to air, which occur during traditional sample preparation and transfer steps. The simple, and therefore extremely reliable, tube

punching mechanism of REMP Tube Technology led to a dramatic decrease of the error rate for tube picking compared to traditional pick and place concepts and this, combined with the well-controlled dry environment (0.4 g water / kg air at -20 °C), made it possible to store dissolved compounds in DMSO for years with minimal deterioration. For the storage of mother solutions or other samples, such as biological material, REMP has developed additional tubes in a 96-well format for volumes of 200/300 µl and recently 800/900 µl. These tubes can be closed either by individual seals or with individual caps if repeated access to an aliquot cannot be avoided. Devices and consumables are available to automatically seal and pierce or cap and decap the tubes and all tube transfers are logged in a database so that tubes cannot get lost during transfers. REMP’s sample processing and storage concepts have traditionally been developed with large facilities in mind but now, with the introduction of the Small Size Store™ (SSS) fully automated sample storage and retrieval system, even smallscale research operations can maximize their sample integrity during storage and increase efficiency of sample logistics. The SSS can run at operating temperatures of, for example, -20 °C, -5 °C and +4 °C, at low relative humidity, and tubes can be cherry-picked at any time, leaving the destination tube racks within the storage environment until instructed otherwise by the user or master control system. Two large service doors on the back of the store offer easy service access to the plates and all robotic components.

Tecan Journal 2/2005

SAMPLE MANAGEMENT

Each SSS unit can be configured to store 1060 REMP 384 Storage Tube Racks, 860 REMP 96 Storage Tube Racks (300) or 500 REMP 96 Storage Tube Racks (900), and, once a laboratory outgrows the capacity of a single unit, up to three units can be linked together. The SSS is fully compatible with the powerful REMP Sample Management Software packages and can be operated as a standalone unit or integrated into a robotic system. This expertise in product integration, such as that seen in the SSS, is a shared attribute of both REMP and Tecan. This ideal will form the basis of a strong combined R&D pipeline for the two companies, which will undoubtedly result in the development of a number of exciting new products. REMP Tube Technologyâ&#x201E;˘ and Small Size Storeâ&#x201E;˘ are trademarks of REMP AG, Oberdiessbach, Switzerland

Various formats of REMP Tubes, sealed or capped

Tube Punching Module (TPM) for troublefree "cherry picking"

This ideal will form the basis of a strong combined R&D pipeline for the two companies, which will undoubtedly result in the development of a number of exciting new products.

Automated Plate Replicator (APR) for mass plate production Tecan Journal 2/2005

A P P L I C AT I O N B I O P H A R M A

Tecan’s Freedom EVO

384R

workstation enables complete automation of a high throughput SNPlex™ genotyping assay in Kiel

OLA-preparation robot with Freedom EVO96R system (Freedom EVO384R for PCR set-up in the same lab not visible)

Professor Stefan Schreiber and his team from the Institute for Clinical Molecular Biology at the Christian-AlbrechtsUniversity in Kiel, Germany, have recently added a Freedom EVO384R workstation to their collection of Tecan systems. The Freedom EVO384R package includes a Tecan Te-MO 384-channel pipettor with a RoMa arm on the established Freedom EVO® liquid handling platform (an upgrade with a LiHa arm is possible). This seventh Tecan workstation in Prof Schreiber’s lab adds to the automated genotyping workflow that they are using to identify genetic changes in a number of diseases, with a specific interest in inflammatory bowel disease (IBD), ulcerative colitis and Crohn’s disease. The Institute is a member of the German National Genome Network, which brings scientific groups together from across the country to research diseases that have a high incidence or result in prolonged suffering and premature death. Prof Schreiber’s laboratory genotypes samples

Tecan Journal 2/2005

using Applied Biosystems SNPlex™ technology and the lab’s daily throughput target of over 350,000 genotypes, which corresponds to 20 x 384 format microplates, has led to a real necessity for as much automation as possible, with unequivocal quality control of the processes involved.

The SNPlex technology requires physical separation of the pre- and post-PCR steps so, at Kiel, each stage is carried out on one of the laboratory’s existing Freedom EVO platforms. These are fitted with Tecan Te-MOTM 96- or 384-channel pipettors, respectively Te-MO 96 and TeMO 384, for accurate dispensing of small

PCR reaction preparation system with Freedom EVO384R (Freedom EVO96R for OLA reaction preparation in the same lab is not visible)

A P P L I C AT I O N B I O P H A R M A

Post-PCR robots with Te-MO 96 and Te-MO 384 back to back

volumes in all the liquid handling steps. The automated SNPlex workflow takes two laboratory days in total and, currently, up to 20 plates in the 384-well format can be prepared. In the pre-PCR laboratory, on day one, a Freedom EVO equipped with a Te-MO 96 combines the OLA master mix with phosphorylated primers and probes, and transfers this to 384-well plates containing the purified, dried gDNA. After the OLA reaction is done on an external thermal cycler, the PCR reaction is set up on a Freedom EVO384R, where the OLA product is added to the PCR master mix. On day two, the plates are transferred to the Freedom EVO with a Te-MO 384 and Tecan Power Washer 384™ (PW 384) plate

washer for the post-PCR stages, which include binding of the PCR products to the SNP plate, isolation of the biotinylated strand, hybridization of the ZipChute® probes, elution of the hybridized probes and setting up of the Applied Biosystems 3730xl DNA Analyzer loading plate. Fixed tips were found to be preferable on the Te-MO 384 to lower the amount of waste produced and increase coefficients of variation and the high quality of washing achieved by the PW 384 has had a major effect on the success of each run. Finally, the data are analyzed and checked for quality control. The Tecan robotic platforms have shown excellent performance in automating the SNPlex technology, with an average call rate of 98% and reproducibility and

accuracy levels reaching over 99.8%. Throughput has increased, quality control is excellent and the whole workflow is overseen by just five technicians. The present daily throughput of the laboratory is 20 plates, or 350,000 genotypes, but this is currently limited by the number of thermal cyclers. Once all bottlenecks have been eliminated, the resoundingly successful combination of SNPlex technology with Tecan’s robotic automation is expected to raise this number to over 50 plates, corresponding to an astounding one million genotypes carried out each day. Applied Biosystems is a registered trademark of Applera Corporation or its subsidaries in the U.S. and/or certain other countries. ZipChute and SNPlex are trademarks of Applera Corporation or its subsidaries in the U.S. and/or certain other countries. Freedom EVO® is a registered trademark of Tecan Group Ltd., Männedorf, Switzerland

Post-PCR system based on Freedom EVO384R, the PW 384 and the adjacent Tecan Plate Hotel Tecan Journal 2/2005

A P P L I C AT I O N B I O P H A R M A

Solving protein structures on a high throughput scale Renaud Vincentelli and Christian Cambillau

Renaud explains the automated dot blot to the students of the “EMBO Practical Course on High Throughput Methods for Protein Production and Crystallization” organized in the lab in July 2005

The Architecture et Fonction des Macromolécules Biologiques (AFMB) laboratory in Marseille is currently collaborating in at least four different large-scale international protein structure determination networks (SPINE, VIZIER, E-MEP, MEPNET). The lab’s Structural Genomics team, headed by Dr C. Cambillau, is playing a key role in these investigations, examining the expression and crystallization of proteins within such diverse organisms as bacteria, viruses including HIV and SARS, and mammals. To date, the team has been working on more than a thousand proteins, of which about 800 are soluble proteins and the remainder membrane proteins (secondary active transporters and G protein-coupled receptors (GPCRs)). The team has now established a high throughput facility at the AFMB for which they have developed strategies and automated methods for almost all aspects of protein production and crystallization1. Renaud Vincentelli is the Methods Development Manager at the AFMB and was responsible for setting up the lab’s high throughput protein production platform. Renaud and the Structural Genomics team continue to develop novel, Tecan Journal 2/2005

automated methods for all their protein characterization procedures on a high throughput scale, including cloning, gene expression and protein purification. The lab is currently using two Tecan systems, the first, a Tecan Genesis® 100/8 platform, is dedicated to protein crystallization, performing micro-droplet crystal screens under 1,236 different conditions using 96-well Greiner plates. These also undergo nano-droplet crystallization screens and the automated procedure developed by the AFMB staff allows crystals to be obtained within just a few hours with around 1 mg of protein, instead of taking several days, vastly improving the throughput capability for any lab using the method2. The second instrument, a Tecan Genesis Freedom® 200 liquid handling system, is equipped with eight needles and a variety of added extras including shakers, a vacuum station and a GENios™ Plus reader. This system is used for the majority of the steps involved in the protein expression studies, including PCR mixes, bacterial transformation, minipreps, protein refolding and gene expression screening. Significantly,

quantitative dot blot procedures have also been automated within this system3,4, where the amount of total and soluble recombinant histidine (His)-tagged proteins expressed in E. coli are detected using chemiluminescence with the GENios Plus. The dot blot sensitivity is currently about 0.5 to 1.0 mg per liter of culture and this can be improved by adding a Nickel 96 assay to the protocol. The dot blot detection is linked to upstream bacterial culture treatment. As a result of the constant refinement and optimization of this application, it takes just a few hours from harvesting the cells (automated) to having the complete protein solubility/expression results and the program can autonomously process 160 independent cultures at a time. With this setup the robot can perform up to 12 dot blot plates per day and, if necessary, by using 384-well plates and adding a 96 head to the setup, this throughput could be increased (for instance, for full library screening with other specific antibodies). This dot blot procedure can be used with other antibodies and its automation and quantification advantages mean it could replace the traditional dot blot protocol.

A P P L I C AT I O N B I O P H A R M A

Expression tests

Soluble

Cloning 6XHis Target

Large scale expression

Refolding screening Insoluble

Structure

Crystallization

Refolding

Soluble

The Genesis Freedom system is also used for protein refolding, again the group has developed a completely automated method5 that aims to overcome many of the weaknesses present in existing methods. This was the first completely automated inclusion body refolding screening procedure to be developed using a 96-well format and the procedure is potentially applicable to any nonmembrane protein. As Renaud explained, he chose Tecan’s Genesis Freedom 200/8 for a number of reasons, “The Tecan Freedom was already proven in the market and we knew that the Tecan service was excellent because we already had the Genesis 100/8 platform. The system’s flexibility is invaluable, both in terms of hardware and software – I can add new components, and I can program the software very quickly to do basically everything I want.” Automation of its laboratory methods has proven to be especially important in the AFMB’s contribution to significant international projects such as Structural Proteomics in Europe (SPINE), which aims to develop technologies to determine the structure of both prokaryotic and eukaryotic proteins. The Structural Genomics team is one of 11 core structural biology labs involved and has solved the crystal structure of key proteins from a variety of major human pathogens. The AFMB also joined the Membrane Proteins Network (MePNet) program in 2002, which focuses on

solving the structures of membranebased receptors, especially important therapeutic targets including the GPCR superfamily. Expression, purification and refolding investigations are already underway on some of these 100 proteins and crystallization will soon follow. More recently, an investigation into the integrated structural genomics of viral enzymes involved in replication (VIZIER) has begun, which aims to collect the largest possible amount of structural information on RNA viruses for exploitation by drug development companies. The AFMB is the co-ordinator of this project, which will involve cloning of 3,000 viral proteins and automated expression screening over the next two to three years. The AFMB also provides Platform RIO, an open facility with a wide range of protein characterization services including bioinformatics, protein production, protein crystallization and structure determination.

References: 1. Vincentelli R, et al. (2003) Medium scale structural genomics: strategies for protein expression and crystallization. Accounts Chem. Res. 36, 165-172 2. Sulzenbacher G, et al. (2002) A medium-throughput crystallization approach. Acta Crystallographica Section D 58: 2109-2115 3. Automation of quantitative dot blot procedures – Tecan application note, in press

Purification The AFMB high throughput pipeline, from cloning to solving protein structures

Renaud Vincentelli, Methods Development Manager

4. Vincentelli R, et al (2005). Automated expression and solubility screening of His-tagged proteins in 96-well format. Analytical Biochemistry (in press) 5. Vincentelli R, Canaan, et al. (2004). High-throughput automated refolding screening of inclusion bodies. Protein Science 13: 2782-2792 For further information please contact: Renaud Vincentelli: Renaud.Vincentelli@afmb.univ-mrs.fr AFMB lab: http://www.afmb.univ-mrs.fr RIO service: http://afmb.cnrsmrs.fr/rio/Plateforme_RIO.html Genesis® Freedom is a registered trademark of Tecan Group Ltd., Männedorf, Switzerland

Tecan Journal 2/2005

L AT E ST P R O D U C T S

Tecan up close – new products in detail Two LiHa arms with fixed or disposable tips

One is nice but two are better – dual arms increase the Freedom EVO®’s liquid handling capabilities

The dual LiHa arms can run in parallel with the RoMa arm and are fully compatible with Tecan’s extensive range of application options, including storage elements, stirrers and incubation, separation or reading devices. This capability is a great asset for a variety of applications, including hit picking, serial dilutions, consecutive sample transfer and other aspects of assay preparation, and can greatly enhance the liquid handling power of any laboratory. Fixed or variable, your choice! – Tecan’s new microplate reader series brings Infinite™ 200 series flexibility

The Freedom EVO with dual LiHa arms

Tecan’s Freedom EVO series of automated workstations now offer unrivalled processing speed and flexibility thanks to a unique feature that allows two LiHa arms to be installed onto the same platform, acting completely independently. Each arm has up to eight variable-spacing channels, for use with disposable or washable tips, that can be adapted to handle high volumes, for example of cell culture media, as well as low volumes of precious reagents or samples.

Tecan’s new series of flexible, upgradeable multi-mode microplate readers combine reagent dispensing with several detection modes in an affordable platform for the complete range of life science applications. The unique modular design and adaptability of the Infinite 200 series provides researchers with a choice of monochromator or filter system and allows them to tailor the instrument’s detection modes for their individual needs. The Infinite 200’s monochromator module provides full flexibility in wavelength without the requirement for optical filters, while the filter modules are highly cost efficient for routine applications at fixed wavelengths. The configurable detection modes allow the Infinite 200 to read fluorescence assays from the top and bottom as well as luminescence or absorbance assays in

Two engineers working with the Infinite 200

The Infinite 200 upgradeable multi-mode microplate reader Tecan Journal 2/2005

6- to 384-well microplates, PCR plates and cuvettes. These new readers are the first in a series of products to be launched by Tecan and are supported by the powerful new i-control software, which allows the user to easily set up protocols to suit their own application. MultiCheck™ QC test plates, Multicheck™ software and IQ OQ documentation are also available for periodic performance checks. Pin point accuracy – Tecan’s laser guided Positioning System guarantees accurate high density pipetting The Tecan Positioning System, Te-PSTM, is a high precision, configurable LiHa arm with a laser guided positioning sensor that guarantees reliable and accurate routine use of high density pipetting formats such as 1536-well plates. The technology allows real-time, in-process position monitoring and fine adjustment of the pipetting tip position, so removing any intolerance introduced by a variety of labware on the table. The Te-PS is available as an upgrade on any Freedom EVO platform, enhancing performance in a range of applications including matrix-assisted laser desorption ionization (MALDI) target spotting, protein crystallization, hit picking, PCR in 384-well plates and microarrays.

C U STO M E R S U P P O RT

Introducing high quality disposable troughs for reagents Tecan disposable troughs for Genesis® and Freedom EVO® series have been redesigned to improve the residual volume. Available in natural (transparent) polypropylene or gray polypropylene for light sensitive reagents, these troughs hold 100 ml volume and are equipped with a useful 10 ml scale on the side. The increased acute bevel angle of the base of each trough reduces the residual reagent volume to only 3.4 ml. The natural polypropylene troughs are DNA and DNase/RNase free, do not contain any PCR inhibitors, are free of ATP and pyrogens, and are sterile – all certified by an independent laboratory. State-of-the-art production processes, integrated quality control procedures and computer controlled manufacturing systems guarantee that the Tecan disposable troughs meet stringent quality requirements. All production steps, from the completion of goods through to packaging, are fully automated, and the tools involved are subjected to frequent checks to maintain consistent quality throughout the production process. Each trough production lot is assigned a distinct batch number to improve quality control tracking.

Tecan troughs on Genesis and Freedom EVO platforms To order the disposable troughs please use the following product numbers: Disposable troughs for reagents (natural); product number 10 613 048 Disposable troughs for reagents (gray); product number 10 613 049 Please note that the gray trough products (10 613 049) are a replacement for an existing trough product, product number 10 613 021, which is being discontinued. Trough natural blister – Polypropylene shrink wrapping containing 6 natural disposable troughs for reagents / part number 10 613 048

Trough gray blister – Polypropylene shrink wrapping containing 6 gray disposable troughs for reagents / part number 10 613 049

Tecan Journal, Customer Magazine of Tecan Trading AG., ISSN 1660-5276 Design: OTM/London www.otmcreate.com Photography: Marc Wetli/Zürich www.wetli.com, Günter Bolzern/Zürich www.bolzern.net, Nadja Athanasiou/Zürich Editor: kdm/UK www.kdm-communications.com Print: DAZ Druckerei Albisrieden AG/Zurich www.daz.ch Address: Tecan Switzerland AG, Marketing Communications, Seestrasse 103, CH-8708 Männedorf, Switzerland, journal@tecan.com, www.tecan.com Tecan Group Ltd. is making all efforts to include accurate and up-to-date information into this publication. Yet, it cannot be ruled out that omissions or errors might have occurred. Therefore, Tecan Group Ltd. cannot 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 any time without notice. All mentioned trademarks are protected by law. For technical details and detailed procedures of the specifications provided in this document please contact your Tecan representative. © 2005, Tecan Trading AG, Switzerland, all rights reserved. This brochure may contain reference to applications and products which are not available in all markets. Please check with your local sales representative. Tecan® is in major countries a registered trademark of Tecan Group Ltd., Männedorf, Switzerland Tecan Journal 2/2005

Tecan Sales Offices

REMP Sales Offices

Tecan R&D and manufacturing site

REMP R&D and manufacturing site

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

REMP AG Weststrasse 12 CH-3672 Oberdiessbach Switzerland T +41 31 770 70 70 F +41 31 770 72 66 info@remp.com www.remp.com

Asia (Pte) Ltd +65 644 41 886 Tecan Sales Austria GmbH +43 62 46 89 33 Tecan Sales International GmbH +43 62 46 89 33 Tecan Benelux B.V.B.A. +32 15 42 13 19 Tecan Benelux B.V.B.A. +31 18 34 48 17 4 Tecan Group Ltd., Beijing Rep. Office +86 10 586 95 936 Tecan Deutschland GmbH +49 79 51 94 170 Tecan France S.A.S. +33 4 72 76 04 80 Tecan Italia S.r.l. +39 02 215 21 28 Tecan Sales International GmbH +43 62 46 89 33 Tecan Japan Co. Ltd +81 42 334 88 55 Tecan Nordic AB +46 31 75 44 000 Tecan Nordic AB, Rep. Office Denmark +45 70 234 450 Tecan Portugal +351 21 000 82 16 Tecan Sales Switzerland AG +41 44 922 89 22 Tecan Iberica Instr. S.L. +34 93 490 01 74 Tecan UK Ltd. +44 11 89 300 300 Tecan US Inc. +1 919 361 5200 REMP AG (Switzerland) +41 31 770 70 70

Tecan Journal 2/2005

REMP (USA) Inc. +1 508 429 2200

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