IP Profiling

Overview

The understanding of protein complex assembly and mapping of protein interactions has rapidly grown in recent years due to significant advances in mass spectrometry. Combined with selective enrichment tools we can now probe with high degree of selectivity and sensitivity protein/protein interactions and the individual networks they represent within a protein complex. Modern unbiased IP profiling approaches using mass spectrometry circumvents the need to preselect an antibody to each co-binding entity and can therefore be used for the analysis of protein complexes and enable the discovery of new functional interactions. We recommend submitting samples in control and test pairs. You can submit the elution or the matrix. For the former the elution volume should be ~50µL and for the latter the matrix should be shipped free of buffers. Our workflow is robust towards most common buffers but if you have any concerns feel free to give us a call with your questions.

Steps Involved

• Sample preparation: SDS-PAGE with 10 band excision, automated in-gel digestion with a Digilab ProGest robot.
• LC-MS/MS: Increments of 10 x 0.5hr LC-MS/MS.
• Data Processing: Protein and PTM identification is performed with Mascot. Protein visualization and validation is performed with Scaffold.
• Report generation: PDF/Excel/Scaffold

The turnaround time is 2-3 weeks.

Deliverables

• Written Report containing details of your samples, our methods and a summary of the data.
• An Excel workbook containing the detailed qualitative and quantitative protein identification data. For control and test submissions a differential analysis is included and a list of enriched proteins is provided.
• A Scaffold file to enable your own deeper data mining.

An example report is available upon request.

A service note with some data outlining some fundamentals of the service is available here.