title: Over 4100 protein identifications from a Xenopus laevis fertilized egg digest using reversed-phase chromatographic prefractionation followed by capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry analysis.
abstract: A tryptic digest generated from Xenopus laevis fertilized embryos was fractionated by RPLC. One set of 30 fractions was analyzed by 100-min CZE-ESI-MS/MS separations (50 h total instrument time), and a second set of 15 fractions was analyzed by 3-h UPLC-ESI-MS/MS separations (45 h total instrument time). CZE-MS/MS produced 70% as many protein IDs (4134 versus 5787) and 60% as many peptide IDs (22 535 versus 36 848) as UPLC-MS/MS with similar instrument time (50 h versus 45 h) but with 50 times smaller total consumed sample amount (1.5 μg versus 75 μg). Surprisingly, CZE generated peaks that were 25% more intense than UPLC for peptides that were identified by both techniques, despite the 50-fold lower loading amount; this high sensitivity reflects the efficient ionization produced by the electrokinetically pumped nanospray interface used in CZE. This report is the first comparison of CZE-MS/MS and UPLC-MS/MS for large-scale eukaryotic proteomic analysis. The numbers of protein and peptide identifications produced by CZE-ESI-MS/MS approach those produced by UPLC-MS/MS, but with nearly two orders of magnitude lower sample amounts.
authors.affiliation: Department of Chemistry, University of Notre Dame, Notre Dame, IN, USA.
authors.forename: Xiaojing
authors.initials: X
authors.lastname: Yan
authors.affiliation: Department of Chemistry, University of Notre Dame, Notre Dame, IN, USA.
authors.forename: Liangliang
authors.initials: L
authors.lastname: Sun
authors.affiliation: Department of Chemistry, University of Notre Dame, Notre Dame, IN, USA.
authors.forename: Guijie
authors.initials: G
authors.lastname: Zhu
authors.affiliation: Department of Chemistry, University of Notre Dame, Notre Dame, IN, USA.
authors.forename: Olivia F
authors.initials: OF
authors.lastname: Cox
authors.affiliation: Department of Chemistry, University of Notre Dame, Notre Dame, IN, USA.
authors.forename: Norman J
authors.initials: NJ
authors.lastname: Dovichi
mesh_terms: D000818:Animals; D056148:Chromatography, Reverse-Phase / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D004625:Embryo, Nonmammalian; D005260:Female; D040901:Proteomics / Q000379:methods; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D029867:Xenopus Proteins / Q000032:analysis* / Q000378:metabolism; D014982:Xenopus laevis / Q000196:embryology; D015053:Zygote / Q000737:chemistry
chemical_list: D029867:Xenopus Proteins
keywords: Bottom-up proteomics; CZE-ESI-MS/MS; Electrokinetically driven sheath flow CZE-MS interface; Technology; UPLC-ESI-MS/MS; Xenopus laevis