section line 2 BioSciences section line 1 VWRBiosciences,more than just a helping hand Proteomics round-up What can we offer? In today s world of discovery, technology is critical to a better understanding of life. VWR BioSciences has formed strategic partnerships with industry-leading Contract Research Organisations, offering a complete portfolio of custom services for our customers' scientific discovery and drug development needs. Each partner has been carefully screened and selected by VWR, providing the highest quality services and eliminating the researchers' need to spend countless hours searching for a reputable firm. Want to know more? Contact our specialists and you could get a gorgeous free coffee mug. See our promotion in the Shop! By using VWR s expertise of working with many suppliers, coupled with our life sciences speciality, we have set up a portfolio of suppliers that have been validated for their capabilities, reliability and security on issues such as intellectual property. We are committed to excellence, ensuring that our customers can have the utmost confidence in the value that each of our partners provides. Whether your work is in Genomics, Proteomics, or Pre-Clinical, GLP studies or non-glp, VWR BioSciences has the solution to drive innovation. The following pages give a roundup of our capabilities in the increasingly important Proteomics field. Around Europe we have a network of highly trained specialists who can work with you and select the partners from the portfolio that can best empower your research see the VWRbioMarke shop for details and a chance to win a coffee mug too! Proteomics is the study of all the proteins present in a cell, tissue or organism at any one time. Since cells are constantly responding to their environment, and the proteins are the workhorses of the cell, the proteome is also changing - reflecting the life of the cell. Proteomics can be divided into three categories, (1) Expression Proteomics with focus on analysis of protein expression using 2D electrophoresis & Mass spectrometry, (2) Structural Proteomics with focus on analysis of protein structure using NMR & X-ray crystallography & (3) Functional Proteomics/Interaction Proteomics with focus on analysis of protein function using affinity purification, Mass spectrometry & Yeast 2 Hybrid Systems. Proteomics is important for several reasons as in many cases it is an incorrect modification of a normal protein that leads to the development of human diseases. Proteins are almost always useful biomarkers for diagnosis of diseases and the targets for nearly all drugs used for treatment are proteins. Protein Analysis Alphalyse Pick N Post The Pick n Post Protein Analysis Service is based on a FedEx principle. It provides you with easy access to cutting edge analysis with preparation of your samples taking less than 10 minutes: 1. Order your Pick n Post kit for free at www.pick-n-post.com 2. Open the kit and follow the instructions on how to prepare and ship your samples for analysis. 3. Use the Air Way bill provided in the kit for easy delivery. VWR International Issue 18 September 2007 31
Analytical services offered through Pick N Post Protein identification - Mass spectrometric analysis and database searching of proteins isolated from 1D/2D gels or pure proteins in solution provide you with protein name, gene identifier, amino acid sequence and molecular weight. Phosphoprotein characterisation - affinity purification of phosphopeptides and mass spectrometric analysis for the detection of phosphopeptides, the peptide amino acid sequence and identification of the phosphorylation site. Glycoprotein characterisation - mass spectrometric analysis and treatment with specific glycosidases provides detection of glycopeptides, the peptide amino acid sequence with glycosylation site and possible carbohydrate structure. 2D-gel protein profiling - mass spectrometric analysis can identity up to 1000 proteins from each 2D-gel. Ship your 2D-gel and an image with spots marked for analysis and Alphalyse provide spot picking and protein identification. The data is provided to you electronically as a 2D-gel image with identification data linked to each spot. Molecular weight determination - mass spectrometric analysis for the molecular weight of your intact protein, peptide or oligonucleotide. Edman sequencing - N-terminal Edman sequencing of 5-25 amino acids from the N-terminal end of the protein or peptide. Amino acid analysis - acid hydrolysis and chromatographic analysis for the each amino acid in your purified protein or peptide. This analysis is useful for protein quantification and purity estimation. Typical applications of the Pick n Post service Recombinant Proteins - do these questions sound familiar? Am I expressing the correct protein? What are these contaminating proteins? Does my protein form multimers? Pick n Post will provide answers to these recombinant protein expression questions. Post-Translational Modifications Mapping and characterisation of proteins post-translational modifications enables the fuller understanding of biological pathways and disease processes. The Pick n Post phosphoprotein and glycoprotein characterisation service will help you learn more. Protein Profiling Protein analysis using 2D gel electrophoresis is often used to characterise the overall protein expression from a given cell type or to characterise a subset of proteins that are expressed under specific conditions and fractionated by defined methods. The Pick n Post 2D gel protein profiling service will identify all your spots! Protein Biomarkers The objective of protein biomarker projects is to identify differentially expressed proteins in two samples - diseased vs. non-diseased, treated vs. non-treated or two different strains from the same species. Cut out differentially expressed proteins from 2D gels and you can get protein identification within 8 days even for weak silver stained spots. 32 VWR International Issue 18 September 2007
BioSciences Proteome Factory The proteomic key technology platform significantly increases the chances to identify new drug targets in emerging disease and biomarkers of disease state. This technology platform offers reliable analyses of all kinds of protein containing samples including body fluids, cells, tissues and plants. The innovative proteomics platform ensures high sensitive and simultaneous analyses of a vast amount of proteins. Pharmaco proteomics facilitates investigation of molecular mode of action and toxicology of drugs and other compounds. In combination with new prefractionation techniques lower abundant proteins can be analysed allowing the identification of new potential targets and biomarkers (e.g. by depletion of albumin and other abundant proteins from plasma or liquor (CSF) samples). from Proteome Factory Proteome analysis by extreme high resolution 2D electrophoresis (separation of up to 10,000 proteins) for: Target identification Biomarker (disease marker) & surrogate marker identification Drug mechanism of action & predictive toxicology Differential protein (expression) pattern analysis Phospho & glyco proteomics Immuno proteomics Plasma & CSF (liquor) proteomics with depletion of high abundant proteins 2DE WORKFLOW PROTEIN SEPARATION POWER OF PROTEOME FACTORY S 2DE GELS Tissue, cells, microorganisms,body fluids or other protein containing samples Sample preparation, depletion of high abundant proteins & prefractionation Protein concentration determination & concentration of protein 2DE separation by 20 to 40 cm isoelectric focusing gel & 30 cm SDS-PAGE Immuno proteomics western blot Digitalization of 2DE gels, differential 2DE image analysis Immuno proteomics western blot Staining by silver or fluorescence How it works... Sample is prepared appropriately for 2DE Depletion of high abundance plasma or liquor proteins (optional) Prefractionation by HPLC Protein separation by 2DE: Extreme high resolution 2D electrophoresis (7 x 8 cm, 23 x 30 cm, 30 x 40 cm, 30 x 60cm) Fluorescence, CBB G-250 or silver staining and digitalisation Differential image analysis using Proteomweaver Report with results, statistical analysis (fold changes, p values of t- test) Protein identification and characterisation, N-terminal sequencing This technique has many great advantages: Excellent target fishing technique High quality due to highly experienced experts in protein and proteome analysis Extreme high separation power of 2DE (up to 10000 protein spots depending on sample and gel size) High-sensitivity protein identification Clients retain ownership of data Fee-for-service or multi-year contracts No minimum order required Quick turn-around times with rush services available Complete confidentiality VWR International Issue 18 September 2007 33
Structural biology Proteros Proteros' competitive advantage is the proven capability of producing high-resolution X-ray data and protein-ligand-complexes using its superior process and proprietary Free Mounting System (FMS ), unique Picodropper technology, and the largest catalogue of ready-touse proteins (Gallery Structures) for analysis of protein-ligand-structures. By obtaining faster and better structural data, Proteros has set new standards in protein structure analysis. offered Gallery Structure projects the largest commercially available collection of crystallised proteins (Gallery Structures) facilitates fast generation of complex structures with your compounds and high resolution diffraction data within 4 to 6 weeks. Proteros is continuously enlarging it's collection of Gallery Structures, which now numbers over fifty with recent additions including PLK1 and PLK4. Full Gallery structure list on request. Gene-to-structure projects a full service, including the design of appropriate constructs, expression and purification of protein through to crystallography, within 3 to 6 months. These service projects include: Knowledge-based design of constructs specifically for protein crystallisation purposes Parallel and high throughput expression, purification and crystallisation of all proteins Collection of high-resolution X-ray data using FMS and synchrotron source as required Analysis of new structure Successive analysis of novel co-crystals of protein-ligand complexes using Picodropper technology Structure optimisation projects speciality X-ray structure analysis in the areas of: Optimisation of crystallographic data using FMS Analysis of complex structures with low affinity and solubility using Picodropper technology Crystallisation of proteins Rapid X-ray data collection using synchrotron sources Protein supply having expertise in producing proteins for crystallisation, Proteros can offer high quality active proteins for drug discovery. Gene to protein projects custom production of novel active proteins. Readymade proteins - collection of proteins available for immediate use. Details available on request. Applications Analysis of complex structures of customer compounds with target proteins Analysis of new protein structures Optimisation of crystallographic data and structure resolution using FMS Analysis of complex structures with low affinity and solubility using Picodropper technology Fragment structure analysis: high throughput analysis of complex structures Data collection using 3rd generation synchrotron sources Supply of ready-to-use premium quality proteins Technology transfer: Development of standard operating procedures (SOPs) and processes for customer s in-house activities in structure analysis and protein production Biosiris As an industry leader in the molecular modelling of biological molecules, Biosiris skill is based on scientific expertise and an in-house technology platform. Consequently, it adds considerable value to the research and development activities of its customers. The in-house technology 34 VWR International Issue 18 September 2007
BioSciences platform allows customisation of projects based on the study of proteins, peptides and membranes with regard to their sequence- and structure-activity relationships. The expertise within Biosiris is such that they are able to solve complex problems. Alongside these customised services, Biosiris has also developed two mass appeal services: Epiros for the prediction of B epitopes and Protein Investigator for the in-depth analysis of protein sequence. Epiros TM Epiros TM is an original method for the prediction of linear B epitopes of optimal selectivity and maximum sensitivity It has been uniquely developed to detect the best epitope that will enable the production of antibodies adapted to the immunoassay technique that you will use on a protein. As Biosiris regards each case to be distinct, it treats each case as such. Thanks to Epiros and the considerable expertise and experience of Biosiris specialists, we have been able to greatly improve upon the accuracy of epitope prediction. Protein Investigator TM Protein Investigator TM is a rapid, accurate and in-depth study of a protein from its sequence only. The service includes the prediction of the structural and functional properties of your target using Biosiris own technology platform and a wide selection of the most relevant and up-to-date techniques available on the market. It will efficiently collect the maximum relevant information available for your protein of interest. This new and unique service gains time, enhances your efficiency, decreases your costs and helps you to take strategic decisions. Advantages Use of up-to-date and relevant techniques completed by proprietary procedures Indication of prediction reliability Correlation between predictions Clear and concise report for Bioinformatics non specialists External and objective point of view from experts Low-cost analysis Protein synthesis and antibody services IBA IBA is a competent research partner in the academic community and one of the foremost providers of expression cloning and protein purification technologies as well as delivery technologies for proteins and nucleic acids. The proprietary protein products and services cover the entire drug discovery process. IBA assists its customers from early discovery to market identification. Cloning precise cloning of the desired gene into either IBA s proprietary bacterial expression vectors with optimal control of expression or yeast expression vectors or baculo virus transfer vectors. Depending on the gene template provided by the customer, three different cloning procedures can be performed: Subcloning using restriction enzymes, Cloning using PCR from plasmid DNA, Cloning using PCR from a cdna bank or genomic DNA. Protein expression in E. coli general procedure uses a Strep-tag and/or 6xHistidine-tag purification of the recombinant protein. Then depending on the method, the expression can either be a 200 ml test expression and purification via standard Strep-Tactin or NI-NTA chromatography allowing the customer to then evaluate the yield of the recombinant protein and to estimate further culture volumes required to obtain the requested amount of protein. Expression in S. cerevisiae the yeast Saccharomyces cerevisiae is a eukaryotic host producing soluble recombinant proteins and introducing post-translational modifications. VWR International Issue 18 September 2007 35
The expression is performed in a protease deficient strain. The expression vector for S. cerevisiae is based on a multi-copy 2 µm based plasmid containing a strong and rapidly induced promoter. Protein purification can be performed using Strep-tag and/or 6xHistidine-tag. Expression in insect cells using baculovirus with Strep-tag and/or 6xHistidine-tag purification of the recombinant protein, IBA is able to express precisely matured, folded and processed recombinant proteins. General procedure consists of transfection of insect cells, generation of recombinant viruses, plaque purification, test expression of 6 recombinant viruses, western blot analysis, estimation of the necessary culture volume to obtain the requested amount of protein, high titre virus stock production, culture of the appropriate scale and protein expression, cell harvesting and lysate preparation, and affinity purification via Strep-Tactin and/or Ni-NTA. BioGenes BioGenes develops custom monoclonal antibodies against different antigens like proteins, peptides, haptens, toxic molecules, or venomous proteins. BioGenes guarantees the success of the development of a monoclonal Antibody against proteins and peptides including at least 3 stable clones positively reacting with the antigen - tested in an ELISA or the service is free of charge. Antibody production in varying scales Search of pairs (capture and detector antibodies) Labelling of antibodies Long-term storage and care of clones BioGenes offers the development of immunoassays for qualitative and quantitative determination of antigens according to the customer s requirements. The immunoassays are designed with the use of micro titre plates and include: Competitive assay for determination of haptens Sandwich-assay for determination of proteins The development of immunoassays involves the following work phases: Advantages BioGenes has improved the mouse hybridoma technology and offers Complete Monoclonal Antibody Development: - 0,2 mg purified protein for the complete project (includes at least 2 clones) - Time frame 3-4 months - Delivery of all stable-positive, IgG producing clones (up to 10 clones) - All project results and clones belong to the customer (clone ownership is always the customer s) Neoclone Founded in 1999, NeoClone, focuses on monoclonal antibodies and has a proprietary process for rapid development of monoclonal antibodies as well as a catalogue of high quality monoclonal antibody reagents to basic transcription proteins and several immunoaffinity products for protein purification. NeoClone offers custom monoclonal antibody development services. The customer retains all ownership of the antibodies and associated cell lines. Pricing is project specific and is staged with each phase of the development process to reduce risk to the customer. Standard projects 36 VWR International Issue 18 September 2007
BioSciences produce up to 18 different monoclonal antibodies per antigen, ensuring a high likelihood of obtaining an antibody suitable for the customer s specific application. Advantages The exclusive production process produces mouse monoclonal antibodies faster, more cost effectively, in higher yields, and with greater cell line stability than hybridoma technology. The technique is appropriate for a variety of antigens, including peptides, and is amenable to novel immunisation strategies. NeoClone also specialises in developing polyol-responsive antibodies, which are especially useful for gentle protein purification under non-denaturing conditions. Thermo Biopolymers Thermo Biopolymers is specialised in custom synthesis of biopolymers, such as oligonucleotides, peptides, Heavy Peptides and polyclonal antibodies. Read more on the next page and see our special offer in Shop! Polyclonal peptide antibody services - Polyclonal antibodies are commonly used in research, diagnostic for protein detection and quantification and can be generated against proteins or peptides. Plus a custom polyclonal peptide antibody service. Advantages compared to protein antibodies Peptide antibodies can be generated directly from the amino acid sequence of a protein (either from or DNA sequence. Specific regions of a protein can be targeted Highly cost effective and time saving Flexible, stable and safe solution peptide antibodies Extensive range of services from standard peptide or high grade phosphorylated peptide immunisation to affinity purification A team of experts always ready to assist you quickly and efficiently from peptide selection to delivery of your product High success rate is guaranteed through an efficient peptide prediction service, high quality custom peptides and proper animal care Antibodies are delivered lyophilised, guaranteeing highest stability, the possibility of adjusting titre and cost effective shipment Extensive documentation throughout the process with ELISA results delivered after each bleed test and with the final product Peptide services a one-stop-shop solution for all peptide synthesis requirements, including peptide that many companies wouldn t attempt. Thermo has established expertise in the chemical synthesis of peptides for research use and is committed to providing a high quality product that meets your requirements. Our Functional Peptides are controlled by Mass Spectrometry and analytical HPLC at no extra cost, thus we can provide products that meet the highest quality standard. A team of expert peptide scientists will support you starting from the design phase of your peptide to the right choice of synthesis strategy, modifications or purity for your assay. Just contact the Thermo Technical Support team and we will help you obtain the best experimental results for your application. Heavy Peptides for relative and absolute protein quantification offering ultimate specificity, accuracy and sensitivity. One fundamental and essential question in proteomics is the quantification of proteins at very low concentrations or in complex protein mixtures. To meet the increasing need for specificity, accuracy and sensitivity, a quantitation technique using stable isotope (13C / 15N) labelled internal peptide standards has been developed. Thermo Scientific one of the best and largest supplier of LC-MS in the world is certainly your best partner for protein quantification. HeavyPeptide AQUA kits are the first kits that offer a complete solution both for absolute and relative quantitation of proteins by Mass Spectrometry (MS). VWR International Issue 18 September 2007 37