Enhanced detergent extraction for analysis of membrane proteomes by two-dimensional gel electrophoresis

Matthew A Churchward¹ , R Hussain Butt¹ , John C Lang¹ , Kimberly K Hsu¹ and Jens R Coorssen¹^,2^,3

¹Dept. of Physiology and Biophysics, University of Calgary Faculty of Medicine, 3330 Hospital Drive NW, Calgary AB, T2N 4N1, CANADA

²Dept. of Biochemistry and Molecular Biology, University of Calgary Faculty of Medicine, 3330 Hospital Drive NW, Calgary AB, T2N 4N1, CANADA

³Hotchkiss Brain Institute, University of Calgary Faculty of Medicine, 3330 Hospital Drive NW, Calgary AB, T2N 4N1, CANADA

author email corresponding author email

Proteome Science 2005, 3:5doi:10.1186/1477-5956-3-5


Published:	7 June 2005

Abstract

Background

The analysis of hydrophobic membrane proteins by two-dimensional gel electrophoresis has long been hampered by the concept of inherent difficulty due to solubility issues. We have optimized extraction protocols by varying the detergent composition of the solubilization buffer with a variety of commercially available non-ionic and zwitterionic detergents and detergent-like phospholipids.

Results

After initial analyses by one-dimensional SDS-PAGE, quantitative two-dimensional analyses of human erythrocyte membranes, mouse liver membranes, and mouse brain membranes, extracted with buffers that included the zwitterionic detergent MEGA 10 (decanoyl-N-methylglucamide) and the zwitterionic lipid LPC (1-lauroyl lysophosphatidylcholine), showed selective improvement over extraction with the common 2-DE detergent CHAPS (3 [(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate). Mixtures of the three detergents showed additive improvements in spot number, density, and resolution. Substantial improvements in the analysis of a brain membrane proteome were observed.

Conclusion

This study demonstrates that an optimized detergent mix, coupled with rigorous sample handling and electrophoretic protocols, enables simple and effective analysis of membrane proteomes using two-dimensional electrophoresis.