Protein/peptide purification by three-well OFFGEL electrophoresis with immobilized ultra narrow pH gradient gels

Abstract

Purification and desalting of protein and peptide samples by three-well OFFGEL electrophoresis with immobilized ultra narrow pH gradient gels is proposed as a fast preparative strategy for proteomics. The gist of this strategy is to separate proteins and peptides according to their isoelectric points and to isolate those of a given pI value equal to the mean pH value of the gel. The present approach has been demonstrated both on protein mixtures and a digested Escherichia coli protein extract. UV-Vis spectroscopy, MALDI-MS, SDS-PAGE and LC-MS/MS were employed for the quantitative and qualitative characterization of the separation results. The electrophoretic methodology has been simulated by finite element methods.