Prefractionation of protein samples prior to two-dimensional electrophoresis

Garry L. Corthals, Mark P. Molloy, Ben R. Herbert, Keith L. Williams*, Andrew A. Gooley

*Corresponding author for this work

Research output: Contribution to journalArticle

77 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)317-323
Number of pages7
JournalElectrophoresis
Volume18
Issue number3-4
DOIs
Publication statusPublished - Mar 1997

Keywords

  • native electrophoresis
  • prefractionation
  • preparative electrophoresis
  • proteome
  • two-dimensional polyacrylamide gel electrophoresis

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