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A turning point in proteome analysis: Sample prefractionation via multicompartment electrolyzers with isoelectric membranes
Author(s) -
Herbert Ben,
Righetti Pier Giorgio
Publication year - 2000
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/1522-2683(200011)21:17<3639::aid-elps3639>3.0.co;2-v
Subject(s) - isoelectric focusing , proteome , chromatography , isoelectric point , chemistry , membrane , sample preparation , immobilized ph gradient , resolution (logic) , coomassie brilliant blue , analytical chemistry (journal) , staining , biochemistry , biology , computer science , artificial intelligence , genetics , enzyme
Sample prefractionation, as obtained via multicompartment electrolyzers with isoelectric membranes, greatly enhanced the load ability, resolution and detection sensitivity of two‐dimensional (2‐D) maps in proteome analysis. This was demonstrated with different samples. In an Escherichia coli total cell extract, analysis by a 2‐D map run in a pH 4—5 gradient showed many more spots when prefractionated, as compared with standard maps available in databases such as SWISS‐2DPAGE. Analysis of human plasma in the pH 3—6 range showed an increase in the number of highly acidic proteins in the fractionated sample compared to whole plasma. With both samples no protein precipitation or smears occurred and much larger sample amounts could be loaded upon prefractionation, so that a large number of spots could be visualized by Coomassie staining, which is fully compatible with subsequent matrix assisted laser desorption/ionization‐time of flight (MALDI‐TOF) analysis.