Innovations for Protein Research. Protein Research. Powerful workflows built on solid science

Transcription

1 Innovations for Protein Research Protein Research Powerful workflows built on solid science

2 Your partner in discovering innovative solutions for quantitative protein and biomarker research Applied Biosystems/MDS Analytical Technologies is a leading provider of mass spectrometry based systems and workflows for protein research. Our tools enable you to dig deeper into the proteome than ever before to identify, characterize, and quantify lower abundant proteins, and our intelligent software helps to filter through the mountains of data to get to the biologically relevant results.

3 Intelligent Tools, Relevant Results Our advanced protein research systems and workflows are optimized to obtain the most information from each sample. Our systems are based on industry leading mass spectrometry platforms, and integrate robust, easy to use nano and micro LC systems, tagging reagent technologies, and innovative software that makes it easier to get high confidence results. Our protein research solutions have been developed to provide the highest level of performance for protein identification, characterization, and quantification. Systems for Protein Research 4800 MALDI TOF/TOF Analyzer QSTAR Elite Hybrid LC/MS/MS System 4000 QTRAP LC/MS/MS System 3200 QTRAP LC/MS/MS System Provides maximum coverage for protein ID Designed for MALDI or LC MALDI workflows Powerful workflows derive the most information from each sample Easy, wizard driven software Highest sensitivity More coverage in less time highest speed of acquisition High sensitivity, dynamic range, and mass accuracy Powerful versatility for many proteomics applications Supports electrospray and MALDI workflows Intelligent data acquisition features High sensitivity hybrid triple quadrupole/linear ion trap system High sensitivity targeted analysis Identification confirmation of single peptide ID s Outstanding qualitative performance Wide dynamic range for quantification Hybrid triple quadrupole/ linear ion trap system Simultaneous qualitative/quantitative measurements Excellent performance/ cost value Optimized for targeted analysis

4 Protein identification Recently introduced publishing guidelines have put more focus on the quality of protein identifications, and the ability to quantify proteins helps to put them into biological context. Applied Biosystems/MDS Analytical Technologies systems and workflows have been developed to provide the highest confidence protein identifications as well as accurate quantification of differentially expressed proteins in a single experiment. More Coverage in Less Time Advances in system hardware, electronics, and software have resulted in LC/MS/MS technology that can provide fast, intelligent data acquisition from even the most complex samples. The QSTAR Elite LC/MS/MS system can scan up to 7 MS/MS per second with highly intelligent acquisition capabilities for acquiring non-redundant data from peptides at all intensity levels, making it the perfect compliment to the fast gradient capabilities of the Tempo nano LC system. ProteinPilot Software with the Paragon search algorithm enables intelligent data processing by providing high confidence protein identifications with a minimum of false positives, and the ability to search for over 150 peptide modifications simultaneously. Greatest Depth of Coverage Combining the separation capabilities of LC with the analysis power of a MALDI TOF/TOF analyzer in an LC MALDI workflow provides the ability to get the maximum coverage from a given sample. The LC MALDI workflow separates the MS/MS analysis from the time constraints of LC, allowing you to take more time to fully investigate any region of a chromatogram. Spotted MALDI plates can also be archived and re-investigated at a later date. The 4800 MALDI TOF/TOF Analyzer provides low attomole sensitivity, easy to use software, and intelligent workflows to obtain more information from each experiment. Fast nanoflow gradients can be run on the Tempo nano LC system and QSTAR Elite system while maintaining typical sequence coverage above 50%. Sequence coverage of BSA (100 fmol on column) was > 70% using a 6 minute run time, including re-equilibration. The identification of 1429 proteins with C.I. >95% (Mascot score >37) from 10,000 cells with a simple one dimensional reversed phase separation clearly shows the power of the LC-MALDI workflow in combination with the 4800 MALDI TOF/TOF Analyzer.

5 Targeted Approach to Confirm Low Confidence ID s The MIDAS workflow, a high sensitivity, high specificity, targeted workflow can definitively confirm, or reject, putative protein identifications that are based on a single peptide. A Multiple Reaction Monitoring (MRM) scan on a QTRAP system can target a specific peptide, and automatically trigger a high sensitivity MS/MS scan to provide confirming sequence information. The QTRAP Systems, using MIDAS workflow, provide the highest sensitivity for targeted protein identification. Quantitative Proteomics All of the Applied Biosystems/MDS Analytical Technologies proteomics platforms fully support ICAT, itraq, and SILAC reagent workflows that enable investigation of protein abundance as well as identification. Advanced system software automates data acquisition, analysis, and reporting, and provides comprehensive viewing tools for interpreting results. Imaging MALDI Mass Spectrometry Imaging (MSI) is an emerging technology that allows you to visualize the spacial distribution of peptides and proteins across a tissue section. The 4800 MALDI TOF/TOF Analyzer is uniquely equipped to get the most out of every sample. The Opti-Beam on-axis Nd:YAG laser system provides unsurpassed sensitivity with a firing rate of 200 Hz. Furthermore, the large format MALDI target plates allow multiple tissue sections to be mounted, and the industry standard BioMAP software provides comprehensive image analysis. The 4800 MALDI TOF/TOF Analyzer can generate images in both linear and reflectron mode with a variety of matrices. The logical requirement of many MSI experiments is to identify the peptides and proteins present. To this end, MSI maps generated in reflectron mode can take advantage of the superior QuanTIS precursor ion selection to get high-quality MS/MS spectra that can be searched against standard databases. The itraq reagents and workflow enable simultaneous identification and quantification of multiple samples. This allows dosing, time course, and pooling studies for large sample sets, candidate biomarker identification and quantification, as well as quantification of target proteins. Mountains of Data to Biological Results Intelligent data acquisition, a novel search algorithm, and advanced data analysis and reporting capabilities result in more meaningful information not just more data. ProteinPilot software includes the new Paragon search algorithm that identifies more proteins while minimizing false positives, as well as the Pro Group algorithm, that improves protein ID accuracy by minimizing the reporting of false protein isoforms. ProteinPilot software also includes the industry standard Mascot search engine. BioMAP software enables MSI maps to be overlaid on optical images of the tissue sample allowing direct correlation. Here, a protein is mapped in coronal rat brain tissue and is color-coded for intensity (red is more intense).

6 Protein characterization Characterizing protein sequences, as well as detecting, mapping, and quantitating post-translational modifications can lead to further understanding of protein functions. De novo Sequencing De novo sequencing derives peptide sequence information from MS/MS fragment ion data, with no dependence on database searches. This provides a means to research novel peptides, identify missed and non-specific cleavages, identify homologous sequences, and determine multiple post translational modifications. The high sensitivity and mass accuracy of the QSTAR Elite LC/MS/MS system, combined with selectable automation levels in BioAnalyst software, make it an excellent tool for de novo sequencing. The QSTAR Elite system features controllable CID (collisionally induced dissociation) and quadrupole-like fragmentation that generate high quality b- and y- ions. Also the system is not limited by the low-mass cutoff observed in most ion traps, providing greater fragmentation coverage and immonium ions for each peptide. YRLLNENNVEWTLVLNENLLFR the Tempo LC system provides unmatched performance for generating peptide maps, and BioAnalyst software matches experimental peaks to theoretical peptide masses, while accounting for modifications, missed and nonconformant cleavages, and disulfide bonds. PTM Discovery Workflow Mass spectrometric analysis of post-translational modifications has traditionally required using combinations of instruments and methods. The introduction of hybrid triple quadrupole/linear ion trap technology and advances in automation software have resulted in the development of an automated PTM discovery workflow that can identify and determine the sites of unknown modifications in complex biological samples in a single run. Using linked scan functions, the workflow selectively monitors for modification specific fragment ions, performs an enhanced resolution scan to determine charge state and monoisotopic mass, then runs an enhanced MS/MS scan to provide identification and sequence information. The unique combination of triple quadrupole and ion trap capabilities in the QTRAP LC/MS/MS systems allows these scan functions to be run in a single automated workflow. The high mass accuracy and resolution of the QSTAR Elite system is critical in the de novo sequencing of large peptides. Shown is the reconstructed spectrum of MS/MS of an ion at (4+) representing a 22 amino acid peptide, highlighting the contiguous y-series. Peptide Mapping Peptide mapping is used to confirm the identification of proteins and to validate protein products, such as recombinant proteins used as therapeutics. High confidence peptide mapping requires high mass accuracy, very reproducible chromatography, and advanced software capabilities to match experimental data to theoretical peptide lists. The QSTAR Elite LC/MS/MS system with The PTM discovery workflow combines a highly specific precursor ion scan (m/z -79) with high sensitivity MS/MS scans to identify modified peptides and determine modification sites results that typically require multiple experiments on multiple instruments.

7 High Sensitivity Targeted Analysis In cases where post-translational modifications, such as phosphorylation, on a specific target protein are being investigated, a more sensitive and specific workflow can be applied. A targeted approach can identify and quantify the phosphorylation status of specific proteins during developmental and physiological processes, as well as quantitate degrees of phosphorylation in isoforms. The MIDAS Workflow targeted analysis takes advantage of the triple quadrupole capabilities of the QTRAP systems and uses an MRM scan to screen for hyphothetical phosphopeptide signatures, then uses the ion trap scanning function to get high quality MS/MS full scan data to identify the peptide and determine the site of modification. The triple quadrupole MRM scan function provides the highest sensitivity MS/MS measurement and multiple MRM scans can be run simultaneously to monitor for multiple predicted phosphopeptides in a known protein sequence. Site Specific Quantitation Once phosphorylation sites are identified, changes in response to extracellular stimuli or intracellular events can be determined by using itraq reagents and the 4000 QTRAP system to quantify site specific changes in phosphorylation on a target protein. This workflow utilizes the same precursor ion scan as the PTM discovery workflow (m/z 79) to selectively detect phosphopeptides, and the MS/MS information provides phosphorylation site assignment in addition to quantitative itraq reagent reporter ions that allow for the determination of differences in site occupancy between phosphorylation sites of proteins. MS/MS analysis of LGpSGPDGAEEIKR (S308) from MAPKAP K1 demonstrates the high quality MS/MS fragmentation to accurately assign the site of phosphorylation, as the y- and b- ion evidence confirm the location on the serine 308. The itraq reagent reporter ions (inset) provide relative quantitation and show differences in occupancy of this phosphorylation between active and inactive forms of the protein. Analyzing Noncovalently- Bound Complexes Large noncovalent complexes are notoriously difficult to analyze by mass spectrometry due to their inherent instability. These large gas phase ions typically have a lower net charge than ions produced from denatured proteins of a similar size, resulting in higher m/z values that are outside of the mass range of most commercial instruments. The QSTAR Elite quadrupole / timeof-flight LC/MS/MS system with the patented IonCooler Guide increases the pressure in the Q0 region of the analyzer to increase collisional cooling of large noncovalent complexes to overcome mass analysis challenges, and increases the sensitivity for larger complexes (>0.5 MDaltons) by ten to one thousand-fold. Intact Protein Analysis To confirm the molecular weight of intact proteins such as antibody therapeutics, tools are required to confidently calculate molecular weights from multiply-charged data. The extended mass range, high mass accuracy and resolution of the QSTAR Elite system complimented by advanced processing tools in BioAnalyst software provide an ideal solution for intact protein MW determination. Sophisticated protein deconvolution and reconstruction tools in BioAnalyst Software provide accurate mass determination with the ability to clearly differentiate components within heterogeneous mixtures including protein contaminants, isoforms, glycoforms, and other post-translational modifications.

8 Leading the way in protein research Applied Biosystems/MDS Analytical Technologies is a leading provider of advanced solutions for protein identification, characterization, and quantification. Our Mass spectrometry platforms, LC systems, reagents, and software are intelligently integrated to enable workflows that allow you to get the most information from each sample. When you choose Applied Biosystems/MDS Analytical Technologies as your partner, you re choosing to work with the leading developer of mass spectrometry technologies. We are committed to remaining at the forefront of life science research by providing you the tools you need to succeed. Support You Can Depend on Whenever and Wherever You Need it Our products are backed by one of the world s most extensive service and support organizations staffed by professionals who are trained and dedicated to keep your system running at peak performance 24 hours a day, seven days a week. Maximize Instrument Uptime with Smart Services With Smart Services, we can remotely and proactively track critical system parameters and identify potential instrument problems before they affect your lab s efficiency. Using secure Internet connections between instruments and Applied Biosystems support center, our award-winning Smart Services are helping many of our customers achieve a significant increase in instrument uptime and improve productivity. For Research Use Only. Not for use in diagnostic procedures. Applied Biosystems, AB (Design) and itraq are registered trademarks of Applied Biosystems or its affiliates in the US and/or certain other countries. QSTAR and QTRAP are registered trademarks and BioAnalyst, IonCooler, Tempo, ProteinPilot, Pro Group, Paragon, MALDI TOF/TOF, MIDAS, Opti-Beam and QuanTIS are trademarks of Applied Biosystems/ MDS Analytical Technologies, a joint venture between Applied Biosystems and MDS Inc. MDS is a registered trademark of MDS Inc. SILAC is a trademark of Invitrogen. ICAT is a registered trademark of the University of Washington and is exclusively licensed to Applied Biosystems. All other trademarks are the sole property of their respective owners. Copyright Applied Biosystems and MDS Inc. All Rights Reserved. Printed in the USA, 02/2009 Publication 114BR23-02 Headquarters 850 Lincoln Center Drive Foster City, CA USA Phone Toll Free International Sales For our office locations please call the division headquarters or refer to our Web site at