CZE-ESI-MS/MS system for analysis of subnanogram amounts of tryptic digests of a cellular homogenate

We report the performance of capillary zone electrophoresis coupled with an electrokinetically pumped electrospray interface and an Orbitrap-Velos mass spectrometer for high sensitivity protein analysis. We first investigated the system for quantitation of the tryptic digest of BSA. The system produced outstanding linearity with respect to peak height, number of peptide IDs, and spectral counts across the range of 12 nM to 750 nM (60 amol to 3.5 fmol) of BSA injected. One peptide produced a detection limit of 0.3 nM (1.5 amol) injected. We also analyzed 700 pg of a tryptic digest prepared from a RAW264.7 cell lysate; ten proteins were identified in triplicate analyses after filtering the data with peptide confidence value as high. This sample size corresponds to the protein content of approximately ten eukaryotic cells.