Trypsin digestion of proteins on intact immobilized pH gradient strips for surface matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometric analysis
- 17 September 2003
- journal article
- research article
- Published by Wiley in Rapid Communications in Mass Spectrometry
- Vol. 17 (20) , 2323-2326
- https://doi.org/10.1002/rcm.1199
Abstract
Isolelectric focusing (IEF) of proteins on immobilized pH gradient (IPG) strips is an integral part of two‐dimensional (2D) electrophoresis‐based proteomics. Proteins can be effectively analyzed by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOFMS) on the intact strip itself, leading to the creation of a virtual 2D map giving pI and MW information, bypassing the second dimension SDS‐PAGE. Further, trypsin digestion of proteins on the strip can significantly aid the identification of IPG‐separated proteins. However, the small size of the peptides leads to diffusion along and outside the gel matrix. In this study, we describe a simple spray‐based procedure to perform ‘on‐strip’ trypsin digestion of proteins embedded in IPG strips. Examination of intact myoglobin and its tryptic peptides shows that post‐digestion diffusion of tryptic peptides is significantly minimized using this approach. Copyright © 2003 John Wiley & Sons, Ltd.Keywords
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