We have developed a robust nanoelectrospray sheath flow interface based on an electrokinetic pump to generate very stable and low sheath-flow (1). This electrospray interface is used to couple capillary zone electrophoresis and capillary isoelectric focusing to an Orbitrap Velos mass spectrometer. We have used this system for the bottom-up analysis of the secretome of M. marinum (2); over 140 proteins were identified in 165 minutes of mass spectrometer time. Recently, we described the extension of this technology to the analysis of the E. coli proteome (3); over 1,300 tryptic peptides were identified in a single 50-minute analysis. We have also coupled an integrated on-column microreactor employing immobilized trypsin at the proximal end of the separation capillary. This system was used to digest and analyze 300 pg of the RAW 264.7 cell lysate (4). Finally, we have identified 37 host-cell proteins in a recombinant human monoclonal antibody using capillary isoelectric focusing (5). Protein-A/protein-L colums were used to deplete the antibody. Four hours were required to digest the depleted sample using imobilized trypsin and to perform capillary isoelectric focusing-tandem mass spectrometry analysis of the digest.(1) Wojcik R, Dada OO, Sadilek M, Dovichi NJ. Rapid Commun Mass Spectrom.2010; 24: 2554-60.(2) Li Y, Champion MM, Sun L, Champion PA, Wojcik R, Dovichi NJ. Anal Chem.2012; 84: 1617-22.(3) Zhu G, Sun L, Yan X, Dovichi NJ. Anal Chem.2013; 85: 2569-73.(4) Sun L, Zhu G, Dovichi NJ. Anal Chem. 2013, DOI:  10.1021/ac400523x.(5) Zhu G, Sun L, Wojcik R, Kernaghan D, McGivney JB 4th, Dovichi NJ. Talanta 2012; 98: 253-6

Capillary electrophoresis using an electrokinetically-pumped nanospray interfacefor bottom-up proteomics

246th National Meeting (2013)

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