Protein Profiling Assays – GeLC-MS 10/40

Application

1. Immunoprecitation Profiling Assay

2. Cell Profiling Assay

3. Plasma Profiling Assay

4. CSF Profiling Assay

5. Tissue Profiling Assay

Features and Benefits

• Comparative: compare observed proteins between samples

• Quantitative: relative quantitation of ob-served proteins between samples

• Sensitive:

- GeLC-MS10: identify up to 1000 proteins

- GeLC-MS40: Plasma >600 proteins, CSF > 900 proteins, Tissue >1500 proteins, Cells > 4000 proteins

• Accurate: very low false identification rates (<0.5%)

• Rapid: turnaround in 2 weeks

• Assay Development: analysis serves as a precursor to potential development of a fully quantita-tive, multiplex assay for ‘discovered’ proteins of interest

Technology

GeLC-MS is a powerful but simple approach for proteomic analyses. Samples are separated using 1-D SDS-PAGE. Post separation the gel lane is divided into equally sized segments. Proteins in each segment are reduced and alkylated, and in-gel digestion performed using a site-specific protease such as trypsin. An LC-MS/MS experiment is then performed to obtain peptide sequence information and hence identify the proteins in each segment. Data are collated in a non-redundant list per gel lane.

The steps involved in a GeLC-MS experiment are:

• Sample preparation
• SDS-PAGE
• Segmentation
• Automated in-gel digestion
• NanoLC/MS/MS
• Data analysis – including Mascot, SEQUEST, and Scaffold
• Statistical analysis to confirm changing proteins

SDS-PAGE Coupled to Liquid Chromatography-Tandem Mass Spectrometry: GeLC/MS

Statistical Significance

We recommend investigating proteins that are statistically significant (p<0.05) and that meet the following criteria: > 2-fold change with >=10 spectral counts. We also recommend additional testing for proteins that are not statistically significant but meet the same criteria. We highly recommend the use of at least 4 controls and 4 effected samples for the most cost effective and statistically powered discovery experiment.

GeLC-MS 40 Workflow Example

Contact us directly referencing Product: GeLC-MS

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