Single-shot proteomics using capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry with production of more than 1250 Escherichia coli peptide identifications in a 50 min separation.

Capillary zone electrophoresis (CZE)-electrospray ionization-tandem mass spectrometry (ESI-MS/MS) was optimized and applied for analysis of 1-100 ng Escherichia coli protein digests in a single run (single-shot analysis). The system employed an electrokinetically pumped nanospray interface, a coated capillary, and stacking conditions for sample injection. More than 1 250 peptides were identified by optimized single-shot CZE-ESI-MS/MS with 100 ng digest loaded and 50 min analysis time. When 10 ng and 1 ng digests were loaded, about 1 000 and 600 peptides were identified in a single-shot analysis, respectively. Compared with single-shot ultraperformance liquid chromatography (UPLC)-ESI-MS/MS, CZE-ESI-MS/MS produced fewer peptide IDs (1 377 ± 128 vs 1 875 ± 32) for large sample loading amounts (100 ng) with the same mass spectrometer time (50 min). However, when the loaded digest was mass limited (1 ng), CZE-ESI-MS/MS generated many more peptide identifications than UPLC-ESI-MS/MS (627 ± 38 vs 342 ± 113). In addition, CZE-ESI-MS/MS and UPLC-ESI -MS/MS provided complementary peptide level identifications. These results suggest that CZE-ESI-MS/MS may be useful for large-scale, comprehensive, and confident proteomics analysis.

[1]  Liangliang Sun,et al.  Comparison of the LTQ-Orbitrap Velos and the Q-Exactive for proteomic analysis of 1-1000 ng RAW 264.7 cell lysate digests. , 2013, Rapid communications in mass spectrometry : RCM.

[2]  J. Jorgenson,et al.  Capillary zone electrophoresis. , 1983, Science.

[3]  J. Yates,et al.  Identification of proteins in complexes by solid-phase microextraction/multistep elution/capillary electrophoresis/tandem mass spectrometry. , 1999, Analytical chemistry.

[4]  Liangliang Sun,et al.  A rapid cIEF-ESI-MS/MS method for host cell protein analysis of a recombinant human monoclonal antibody. , 2012, Talanta.

[5]  M. Hearn,et al.  Optimization of field-amplified sample injection for analysis of peptides by capillary electrophoresis-mass spectrometry. , 2006, Analytical chemistry.

[6]  M. Moini.  Simplifying CE-MS operation. 2. Interfacing low-flow separation techniques to mass spectrometry using a porous tip. , 2007, Analytical chemistry.

[7]  M. Mann,et al.  Higher-energy C-trap dissociation for peptide modification analysis , 2007, Nature Methods.

[8]  Rawi Ramautar,et al.  High capacity capillary electrophoresis-electrospray ionization mass spectrometry: coupling a porous sheathless interface with transient-isotachophoresis. , 2010, Analytical chemistry.

[9]  N. Dovichi,et al.  On-line amino acid-based capillary isoelectric focusing-ESI-MS/MS for protein digests analysis. , 2012, Analytica chimica acta.

[10]  J. Yates,et al.  Improving the comprehensiveness and sensitivity of sheathless capillary electrophoresis-tandem mass spectrometry for proteomic analysis. , 2012, Analytical chemistry.

[11]  Steven P Gygi,et al.  Target-decoy search strategy for increased confidence in large-scale protein identifications by mass spectrometry , 2007, Nature Methods.

[12]  M. MacCoss,et al.  Capillary electrophoresis with Orbitrap-Velos mass spectrometry detection. , 2012, Talanta.

[13]  E. Maxwell,et al.  Twenty years of interface development for capillary electrophoresis-electrospray ionization-mass spectrometry. , 2008, Analytica chimica acta.

[14]  Z. Shihabi,et al.  Stacking in capillary zone electrophoresis. , 2000, Journal of chromatography. A.

[15]  N. Dovichi,et al.  Quantitative multiple reaction monitoring of peptide abundance introduced via a capillary zone electrophoresis-electrospray interface. , 2012, Analytical chemistry.

[16]  R. Ramautar,et al.  CE-MS for proteomics: Advances in interface development and application. , 2012, Journal of proteomics.

[17]  N. Dovichi,et al.  CZE‐ESI‐MS/MS system for analysis of subnanogram amounts of tryptic digests of a cellular homogenate , 2012, Proteomics.

[18]  B. Sarg,et al.  Optimization and evaluation of a sheathless capillary electrophoresis-electrospray ionization mass spectrometry platform for peptide analysis: comparison to liquid chromatography-electrospray ionization mass spectrometry. , 2011, Analytical chemistry.

[19]  M. Mann,et al.  Feasibility of large-scale phosphoproteomics with higher energy collisional dissociation fragmentation. , 2010, Journal of proteome research.

[20]  N. Dovichi,et al.  Simplified capillary electrophoresis nanospray sheath-flow interface for high efficiency and sensitive peptide analysis. , 2010, Rapid communications in mass spectrometry : RCM.

[21]  N. Dovichi,et al.  Capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry as an alternative proteomics platform to ultraperformance liquid chromatography-electrospray ionization-tandem mass spectrometry for samples of intermediate complexity. , 2012, Analytical chemistry.

[22]  Alexey I Nesvizhskii,et al.  Empirical statistical model to estimate the accuracy of peptide identifications made by MS/MS and database search. , 2002, Analytical chemistry.