TOWARDS STOICHIOMETRIC SILVER STAINING OF PROTEINS RESOLVED IN COMPLEX 2-DIMENSIONAL ELECTROPHORESIS GELS - REAL-TIME ANALYSIS OF PATTERN DEVELOPMENT

Toward the ultimate goal of deriving quantitative protein data from silver-stained proteins resolved in complex two-dimensional electrophoresis gels we have performed computerized real-time analysis during pattern protein ''silver stain'' development. Three points emerged from this study: (i) Development time is a far more important variable than originally envisioned. (ii) Previous studies which, on the basis of a single development time, sought to relate silver stain slope and threshold properties to amino acid composition are non-informative since it will be shown that slope values change markedly with development time. (iii) The combination of real-time analysis and total protein load consideration allows, for the first time, protein ''spots'' to be grouped into categories, which in turn aid in quantification.