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  • Title: Trypsin digestion of proteins on intact immobilized pH gradient strips for surface matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analysis.
    Author: Iyer S, Olivares J.
    Journal: Rapid Commun Mass Spectrom; 2003; 17(20):2323-6. PubMed ID: 14558133.
    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.
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