Proteomics & Metabolomics Shared Resource

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Equipment 4800 MALDI-TOF-TOF Mass Spectrometer

4800 MALDI-TOF-TOF Mass Spectrometer

PMSR previously housed a 4700 instrument, which was upgraded to 4800 Proteomics Analyzer in 2007. With its superior optics, user friendly software workflows, and improved MS/MS performance, the 4800 system enables confident, high-productivity protein identification and discovery applications. The 4800 MALDI TOF/TOF analyzer has high mass accuracy and is interfaced with the MASCOT search engine for protein identification and data analysis.

QSTAR Elite Hybrid Nano UPLC-MS/MS System

QStar Elite Hybrid Nano UPLC-MS/MS System

The QSTAR Elite System is a hybrid quadrupole time of flight system with an electrospray ionizing source making it a very efficient system for the identification of low level proteins and metabolites in biological samples. Using this instrument we can accurately detect and quantify up- and down regulated proteins and small molecules in complex biological samples with a linear dynamic range of up to four orders of magnitude. The QSTAR is interfaced with the UPLC system (Waters Corp.) which is a high pressure, high sensitivity and high resolution protein separation system. PMSR has used it successfully for various proteomic applications with high run to run reproducibility. The analyst software is used to acquire data using the QSTAR Elite. The raw data is processed by the Protein Pilot (version2) software which uses the MASCOT or 'Paragon' as the database search engine for protein identification purposes.

Tempo Nano MDLC System

Tempo Nano MDLC System

The Tempo Nano MDLC System is used in conjunction with the Probot MALDI spotter (Dionex) system provides an efficient protein separation system for further analysis using the 4800 Proteomics Analyzer. The LC-MALDI service offered by the PMSR provides a very useful tool to researchers, particularly for serum based peptide biomarker discovery experiments. It can also be used with the same ease and efficiency for resolution of complex protein mixtures from a variety of samples.

QTOF Premier Mass Spectrometer Connected to Acquity UPLC System

QTOF Premier Mass Spectrometer

The Waters Micromass Q-TOF Premier Mass Spectrometer is a hybrid quadrupole, orthogonal acceleration time-of-flight mass spectrometer that enables automated exact mass measurement of precursor and fragment ions to yield the high confidence for metabolite profiling projects. The Water's Acquity Ultra performance liquid chromatography system which serves as the front end to the QTOF-MS provides a very efficient, reproducible and a high-throughput system for large scale analysis of samples. This system has been in use since January 2008 and has been a workhorse for the high through put metabolomic experiments. PMSR has successfully tested the robustness of this system by acquiring data for serum samples from a cohort of 1556 patients in one run, lasting 11 days. The chromatography was extremely reproducible with minimal peak drift and the mass accuracy remained within 8ppm for the standards that were run after every hundred injections for the purposes of performance and quality control testing.

4000 QTrap Mass Spectrometer Connected to Nano/ Regular UPLC System

4000 QTrap Mass Spectrometer

The 4000 Q TRAP LC/MS/MS System is a hybrid triple quadrupole/linear ion trap mass spectrometer. It is a very useful addition to the PMSR because it is suited for a wide range of applications and has a broad mass range for analysis of a given sample. This is a recent addition to the PMSR and will be primarily used for drug metabolism studies, metabolite identification and proteomic applications including post-translationally modified protein discovery, protein identification, and biomarker validation. It is online with the ultra performance LC (UPLC) system which is designed for nano-scale as well as regular scale, capillary, and narrow-bore separations to attain the highest chromatographic resolution, sensitivity, and reproducibility. We have used this instrument to validate acetylation sites in 'SUMO' which were detected with the QSTAR Elite LCMS/MS system.

2D Electrophoresis and Analysis

2D Electrophoresis

The PMSR is equipped for 2D gel electrophoresis apparatus with an IEF cell (Bio-Rad, USA), the DALT6 large format 2D electrophoresis system (Amersham Biosciences), Dymension software (Syngene) for 2D gel image analysis. The PMSR uses DeCyder Extended Data Analysis Software (DeCyder EDA) for the analysis of 2D gels for the DIGE workflow. Differential electrophoresis (2-D DIGE) includes an internal standard for every spot on every 2-D gel to give the most accurate, reproducible and dependable quantitation possible. DeCyder 2-D Differential Analysis Software significantly increases throughput by accurately addressing measurement of protein differences with statistical confidence and reduces hands-on time from days to minutes with minimal user-to-user variation. The software efficiently detects, matches, and analyzes protein spots in multiplexed fluorescent images, and is able to detect small differences between protein spots with high statistical confidence.

DIGE Imager and Spot Picker

Dige Imager

The Ettan DIGE Imager is a scanning CCD camera, designed for a variety of applications. By combining high resolution with precise motion control, the Ettan DIGE Imager produces accurate multi channel images of Cy2, Cy3, and Cy5 labeled gels. The system can also image a wide range of other fluorescent gel applications because of high sensitivity for imaging faint spots. The spot picker (GE Healthcare) can pick and transfer six spots a minute. Picking is totally separated from image acquisition and image analysis; therefore, neither technique is compromised. It is optimized with DeCyder software output for DIGE analysis.

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