title: Capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry for quantitative parallel reaction monitoring of peptide abundance and single-shot proteomic analysis of a human cell line.
abstract: We coupled capillary zone electrophoresis (CZE) with an ultrasensitive electrokinetically pumped nanospray ionization source for tandem mass spectrometry (MS/MS) analysis of complex proteomes. We first used the system for the parallel reaction monitoring (PRM) analysis of angiotensin II spiked in 0.45mg/mL of bovine serum albumin (BSA) digest. A calibration curve was generated between the loading amount of angiotensin II and intensity of angiotensin II fragment ions. CZE-PRM generated a linear calibration curve across over 4.5 orders of magnitude dynamic range corresponding to angiotensin II loading amount from 2amole to 150fmole. The relative standard deviations (RSDs) of migration time were <4% and the RSDs of fragment ion intensity were ∼20% or less except 150fmole angiotensin II loading amount data (∼36% RSD). We further applied the system for the first bottom up proteomic analysis of a human cell line using CZE-MS/MS. We generated 283 protein identifications from a 1h long, single-shot CZE MS/MS analysis of the MCF7 breast cancer cell line digest, corresponding to ∼80ng loading amount. The MCF7 digest was fractionated using a C18 solid phase extraction column; single-shot analysis of a single fraction resulted in 468 protein identifications, which is by far the largest number of protein identifications reported for a mammalian proteomic sample using CZE.
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556, USA.
authors.forename: Liangliang
authors.initials: L
authors.lastname: Sun
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556, USA.
authors.forename: Guijie
authors.initials: G
authors.lastname: Zhu
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556, USA.
authors.forename: Si
authors.initials: S
authors.lastname: Mou
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556, USA.
authors.forename: Yimeng
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556, USA.
authors.forename: Matthew M
authors.initials: MM
authors.lastname: Champion
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556, USA. Electronic address: ndovichi@nd.edu.
authors.forename: Norman J
authors.initials: NJ
authors.lastname: Dovichi
mesh_terms: D000818:Animals; D002417:Cattle; D002460:Cell Line; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D010455:Peptides / Q000737:chemistry* / Q000302:isolation & purification; D011506:Proteins / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D012710:Serum Albumin, Bovine / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D010455:Peptides; D011506:Proteins; D012710:Serum Albumin, Bovine
keywords: CZE-ESI-MS/MS; CZE-PRM; Human cell lysate digest; Shotgun proteomics; Ultrasensitive electrokinetically pumped sheath flow nanospray interface
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