Automated analysis of mouse serum peptidome using restricted access media and nanoliquid chromatography-tandem mass spectrometry.

We demonstrate use of restricted access media with reversed phase functionality (RAM-RP) for analysis of low molecular weight proteins and peptides in mouse serum (75 μl) using a custom designed modular automated processing system (MAPS). RAM-RP fractionation with simultaneous removal of high molecular weight and high abundance proteins is integrated with a follow-on buffer exchange module (BE) to ensure compatibility with subsequent processing steps (trypsin digestion and intact peptide separation prior to mass spectrometric analysis). The high sample capacity afforded by chromatographic methods generates enough sample to achieve comprehensive serum peptidome identification (357 proteins) through tandem mass spectrometric analysis of both intact and digested peptides.