Abstract
Staining protocols for PAGE have to be sensitive and should not impair further MS analysis of selected samples. In this study, the MS compatibility of different silver- and Coomassie-staining protocols with a nano-LC-MS/MS system was systematically elucidated. Altogether, 13 different silver-staining, 1 imidazole-staining and 2 Coomassie-staining protocols were used and compared to each other for their achieved sequence coverage and their detection sensitivity. Three proteins were used as model proteins (bovine serum albumin, rabbit L-lactate dehydrogenase, bovine milk beta-lactoglobulin) in decreasing concentration (12 pmol down to 30 fmol) and different staining protocols were applied. The conclusion of this study is that two silver-staining protocols (Blum, H. et al.,. Electrophoresis 1987, 8, 93-99 and Shevchenko, A. et al.,. Anal. Chem. 1996, 68, 850-858) combine good sequence coverage and good sensitivity and are recommended for nano-LC-MS/MS analysis.
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