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IP-works

Introduction

The MS Bioworks IP-works service is designed for the analysis of immunoprecipitated or co-immunoprecipitated proteins. The workflow underlying our service was designed to enable routine analysis of samples generated by immunoprecipitated proteins. As such IP-works is a robust and in our hands routine method for analyzing protein-protein interactions.

Background

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. A classical approach to the analysis of protein-protein interactions may involve SDS-PAGE and Western blot to test a hypothesis and validate a putative interaction. However this approach can leave an investigator blind to novel co-ligands of interest since an antibody must be preselected for Western analysis. 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.

Our approach

Our LC-MS/MS workflow generates a non-redundant list of proteins present in your submitted sample; when two or more samples are being compared a list of putative interaction partners is provided based on clearly defined criteria. Reports are simple and comprise of a PDF document documenting your samples and our methods and a summary of the results. An Excel workbook is provided that includes all of the qualitative and quantitative protein mass spectrometry data. We also provide a Scaffold file for each project, the Scaffold file viewer is free and can be used to view data in more depth than in captured in the Excel workbook e.g. you can view individual product ion spectra and you can look at the sequence coverage obtained for each identified protein. You can download an example report from our FTP server and if you would like help using Scaffold our experts are on hand, please call 734-929-5083 or email for details.

What next?

Questions are good and we welcome them. The most frequent questions we receive are regarding elution buffers, elution volumes, suitable controls, how to interpret data, the availability of databases for unusual organism, compatibility with transcriptomics and the compatibility of our methods with your enrichment mechanism. We take great pride in educating and learning from our clients. Call 734.929.5083 or email and we’ll help.

Success

References are available on request and if you would like to see some recent published examples of the IP-works service please check out our publication page. Some research highlights are listed below. If you use data from MS Bioworks IP-works service please let us know and we will add your paper to the list.

Temporal Association of Herpes Simplex Virus ICP4 with Cellular Complexes Functioning at Multiple Steps in PolII Transcription
Wagner LM, Deluca NA
PLoS One, 2013, Oct 11;8(10):e78242. doi: 10.1371/journal.pone.0078242
http://www.ncbi.nlm.nih.gov/pubmed/24147125

TET2 and TET3 regulate GlcNAcylation and H3K4 methylation through OGT and SET1/COMPASS
Rachel Deplus, Benjamin Delatte, Marie K Schwinn, Matthieu Defrance, Jacqui Mendez, Nancy Murphy, Mark A. Dawson, Michael Volkmar, Pascale Putmans, Emile Calonne, Alan H Shih, Ross L Levine, Olivier Bernard, Thomas Mercher, Eric Solary, Marjeta Urh, Danette L Daniels and Francois Fuks.
The EMBO Journal 2013, 1-11
http://www.ncbi.nlm.nih.gov/pubmed/23353889

A Conserved motif in the C-terminal of DNA polymerase α tether primase to the eukaryotic replisome
M.L. Kilkenny, G. De Piccoli, R.L. Perera, L. Pellegrini
Journal of Biological Chemistry, 2012, 287, 23740-23747
http://www.ncbi.nlm.nih.gov/pubmed/22593576