Micro-proteome analysis using micro-chromatofocusing in intact protein separations.
Multi-dimensional liquid-based separation is required for fractionation and mapping of complex protein mixtures from cells. A method that has been used as the first dimension in such separations is chromatofocusing (CF), which is based on generating a pH gradient on an anion exchange column. The use of pH in the first dimension is essential where pH is a fundamental property of proteins and can provide information on post-translationally modified forms of a protein. In this work, a micro-chromatofocusing technique is introduced which can separate microgram levels of proteins from cell lysates for further analysis by LC-MS/MS. It is shown that this method can analyze 10 microg of sample and detect nearly 700-800 proteins from ovarian cancer cell line lysates.
Kim
Hyeyeung
H
Department of Chemistry, University of Michigan, Ann Arbor, MI 48109, USA.
Lubman
David M
DM
D002853:Chromatography, Liquid / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D036103:Nanotechnology; D011506:Proteins / Q000302:isolation & purification*; D020543:Proteome; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods
D011506:Proteins; D020543:Proteome