Nano-flow multidimensional liquid platform using strong ion-exchange and reversed-phase chromatography for improved proteomic analyses

In this study, an integrated multidimensional nano-flow liquid chromatography platform was established with the combination of peptide separation via online SCX, and successfully applied for proteome analysis. By this platform, peptides were first separated by a SCX microcolumn under a series of salt steps, separated by nano-reversed phase liquid chromatography, and finally identified by ESI-MS/MS. To evaluate the performance, the rat hippocampal cytosol proteins were analyzed. Ten fractions were loaded onto a SCX column via autosampler and analyzed by LC-MS/MS. More than 3323 spectra matched to peptides, and 3141 unique peptide were searched. A total of 600 proteins were identified in this single analysis. Compared with the results obtained by offline 2D-Nano-RPLC-MS/MS approach, the identified protein number was increased upto 13%, with decreased consumed time from 96 to 18 h. These results shows that such an integrated approach would gain attention for better proteome results.