{Reference Type}: Comparative Study
{Title}: Increased proteome coverage by combining PAGE and peptide isoelectric focusing: comparative study of gel-based separation approaches.
{Author}: Atanassov I;Urlaub H;
{Journal}: Proteomics
{Volume}: 13
{Issue}: 20
{Year}: Oct 2013
{Factor}: 5.393
{DOI}: 10.1002/pmic.201300035
{Abstract}: The in-depth analysis of complex proteome samples requires fractionation of the sample into subsamples prior to LC-MS/MS in shotgun proteomics experiments. We have established a 3D workflow for shotgun proteomics that relies on protein separation by 1D PAGE, gel fractionation, trypsin digestion, and peptide separation by in-gel IEF, prior to RP-HPLC-MS/MS. Our results show that applying peptide IEF can significantly increase the number of proteins identified from PAGE subfractionation. This method delivers deeper proteome coverage and provides a large degree of flexibility in experimentally approaching highly complex mixtures by still relying on protein separation according to molecular weight in the first dimension.