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.
Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556, USA.
Liangliang
L
Sun
Guijie
G
Zhu
Yihan
Y
Li
Roza
R
Wojcik
Ping
P
Yang
Norman J
NJ
Dovichi
D000818:Animals; D002417:Cattle; D002457:Cell Extracts / Q000737:chemistry*; D002460:Cell Line; D030562:Databases, Protein; D019075:Electrophoresis, Capillary / Q000379:methods*; D057230:Limit of Detection; D016014:Linear Models; D051379:Mice; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping; D015203:Reproducibility of Results; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014357:Trypsin / Q000737:chemistry* / Q000378:metabolism
D002457:Cell Extracts; D010446:Peptide Fragments; D012710:Serum Albumin, Bovine; D014357:Trypsin