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Prefractionation of protein samples for proteome analysis using reversed-phase high-performance liquid chromatography

Item Type:Article
Title:Prefractionation of protein samples for proteome analysis using reversed-phase high-performance liquid chromatography
Creators Name:Badock, V. and Steinhusen, U. and Bommert, K. and Otto, A.
Abstract:We describe an approach for fractionating complex protein samples prior to two-dimensional gel electrophoresis using reversed-phase high-performance liquid chromatography. Whole lysates of cells and tissue were prefractionated by reversed-phase chromatography and elution with a five-step gradient of increasing acetonitrile concentrations. The proteins obtained at each step were subsequently separated by high-resolution two-dimensional gel electrophoresis (2-DE). The reproducibility of this prefractionation technique proved to be optimal for comparing 2-DE gels from two different cell states. In addition, this method is suitable for enriching low-abundance proteins barely detectable by silver staining to amounts that can be detected by Coomassie blue and further analyzed by mass spectrometry.
Keywords:Low-Abundance Proteins, Prefraction, Proteome Analysis, Reversed-phase Liquid Chromatography, Two-Dimensional Gel Electrophoresis, Animals, Rats
Source:Electrophoresis
ISSN:0173-0835
Publisher:Wiley (Germany)
Volume:22
Number:14
Page Range:2856-2864
Date:1 January 2001
Official Publication:https://doi.org/10.1002/1522-2683(200108)22:14<2856::AID-ELPS2856>3.0.CO;2-U
PubMed:View item in PubMed

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