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Evaluation of several two‐dimensional gel electrophoresis techniques in cardiac proteomics
Author(s) -
Li Zhao Bo,
Flint Paul W.,
Boluyt Marvin O.
Publication year - 2005
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.200500104
Subject(s) - proteome , proteomics , isoelectric focusing , isoelectric point , two dimensional gel electrophoresis , fractionation , gel electrophoresis , chromatography , biology , computational biology , chemistry , biochemistry , enzyme , gene
Two‐dimensional gel electrophoresis (2‐DE) is currently the best method for separating complex mixtures of proteins, and its use is gradually becoming more common in cardiac proteome analysis. A number of variations in basic 2‐DE have emerged, but their usefulness in analyzing cardiac tissue has not been evaluated. The purpose of the present study was to systematically evaluate the capabilities and limitations of several 2‐DE techniques for separating proteins from rat heart tissue. Immobilized pH gradient strips of various pH ranges, parameters of protein loading and staining, subcellular fractionation, and detection of phosphorylated proteins were studied. The results provide guidance for proteome analysis of cardiac and other tissues in terms of selection of the isoelectric point separating window for cardiac proteins, accurate quantitation of cardiac protein abundance, stabilization of technical variation, reduction of sample complexity, enrichment of low‐abundant proteins, and detection of phosphorylated proteins.