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Rapid and effective focusing in a carrier ampholyte solution isoelectric focusing system: a proteome prefractionation tool

Rapid and effective focusing in a carrier ampholyte solution isoelectric focusing system: a proteome prefractionation tool

Journal of Proteome Research 7(4): 1761-1766

Preparative (solution) isoelectric focusing (slEF) is a proven technique for proteome prefractionation, but carries limitations which include the risk of protein loss from isoelectric precipitation, poor focusing, and excessively long separation times. This report describes a simple yet effective method to achieve rapid focusing (as fast as 1 h) and maximize protein recovery using a carrier ampholyte slEF system. Cathodic drift was not present over the time course of the experiment using our eight-chamber device, and we demonstrate the effectiveness of this device for focusing proteome mixtures. We also discuss an MS-compatible acidic wash protocol, which is shown to enhance the recovery of proteins following slEF, thus, improving detection by LC-MS/MS. These approaches overcome significant shortcomings of the technique, enabling effective prefractionation prior to MS analysis.

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Accession: 055361510

Download citation: RISBibTeXText

PMID: 18284188

DOI: 10.1021/pr700677u

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