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Prefractionation of protein samples prior to two‐dimensional electrophoresis
Author(s) -
Corthals Garry L.,
Molloy Mark P.,
Herbert Ben R.,
Williams Keith L.,
Gooley Andrew A.
Publication year - 1997
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/elps.1150180304
Subject(s) - electrophoresis , chromatography , electrokinetic phenomena , fractionation , chemistry , gel electrophoresis of proteins , polyacrylamide gel electrophoresis , membrane protein , biochemistry , membrane , enzyme
Thousands of proteins may be visualised on a two‐dimensional (2‐D) gel, but only hundreds are present at levels sufficient for chemical analysis. Therefore, prefractionation of protein samples prior to 2‐D polyacrylamide gel electrophoresis (PAGE) will be important for the investigation of proteins that are present at sub‐picogram levels in physiological samples. We describe an approach to prefractionate protein samples prior to 2‐D PAGE using the Gradiflow, which is a new (preparative) electrokinetic membrane apparatus designed to fractionate proteins in a number of different ways. We have fractionated human serum under nonreducing conditions using the ‘reflux’ mode, in which proteins are fractionated according to their relative mobility under controlled electrophoretic conditions, where the current is periodically reversed. We describe how fractionation occurs and present examples of enrichment of specific proteins.