Strategy for surveying the proteome using affinity proteomics and mass spectrometry.

Summary, in English

Antibody-based microarrays is a rapidly evolving technology that has gone from the first proof-of-concept studies to more demanding proteome profiling applications, during the last years. Miniaturized microarrays can be printed with large number of antibodies harbouring predetermined specificities, capable of targeting high- as well as low-abundant analytes in complex, nonfractionated proteomes. Consequently, the resolution of such proteome profiling efforts correlate directly to the number of antibodies included, which today is a key limiting factor. To overcome this bottleneck and to be able to perform in-depth global proteome surveys, we propose to interface affinity proteomics with MS-based read-out, as outlined in this technical perspective. Briefly, we have defined a range of peptide motifs, each motif being present in 5-100 different proteins. In this manner, 100 antibodies, binding 100 different motifs commonly distributed among different proteins, would potentially target a protein cluster of 10(4) individual molecules, i.e. around 50% of the nonredundant human proteome. Notably, these motif-specific antibodies would be directly applicable to any proteome in a specie independent manner and not biased towards abundant proteins or certain protein classes. The biological sample is digested, exposed to these immobilized antibodies, whereby motif-containing peptides are specifically captured, enriched and subsequently detected and identified using MS.

• Author: Christer Wingren; Peter James; Carl Borrebaeck
• Department/s: Department of Immunotechnology
• Publishing year: 2009
• Language: English
• Pages: 1511-1517
• Publication/Series: Proteomics
• Volume: 9
• Issue: 6
• Links: http://dx.doi.org/10.1002/pmic.200800802
• Document type: Journal article
• Publisher: John Wiley & Sons Inc.
• Topic: Immunology in the medical area
• Status: Published
• ISBN/ISSN/Other: ISSN: 1615-9861