title: Analysis of the bovine plasma proteome by matrix-assisted laser desorption/ionisation time-of-flight tandem mass spectrometry.
abstract: In this study, the bovine plasma proteome was analysed using a three step protocol: (1) plasma was treated with a combinatorial peptide ligand library (CPLL) to assimilate the differences in concentrations of different proteins in raw plasma; (2) CPLL-treated material was fractionated by three standard electrophoretic separation techniques, and (3) samples were analysed by nano-liquid chromatography (nLC) matrix-assisted laser desorption/ionisation (MALDI) time-of-flight tandem (TOF/TOF) mass spectrometry. The efficiencies of three fractionation protocols for plasma proteome analysis were compared. After size fractionation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), resolution of proteins was better and yields of identified proteins were higher than after charge-based fractionation by preparative gel-free isoelectric focussing. For proteins with isoelectric points >6 and molecular weights ⩾ 63 kDa, the best results were obtained with a 'shotgun' approach, in which the CPLL-treated plasma was digested and the peptides, rather than the proteins, were fractionated by gel-free isoelectric focussing. However, the three fractionation techniques were largely complementary, since only about one-third of the proteome was identified by each approach.
authors.affiliation: Institute of Molecular Biology, Friedrich-Loeffler-Institut, Südufer 10, 17493 Greifswald-Insel Riems, Germany; Institute of Novel and Emerging Infectious Diseases, Friedrich-Loeffler-Institut, Südufer 10, 17493 Greifswald-Insel Riems, Germany.
authors.forename: Ann-Kristin
authors.initials: AK
authors.lastname: Henning
authors.affiliation: Institute of Novel and Emerging Infectious Diseases, Friedrich-Loeffler-Institut, Südufer 10, 17493 Greifswald-Insel Riems, Germany.
authors.forename: Martin H
authors.initials: MH
authors.lastname: Groschup
authors.affiliation: Institute of Molecular Biology, Friedrich-Loeffler-Institut, Südufer 10, 17493 Greifswald-Insel Riems, Germany.
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Mettenleiter
authors.affiliation: Institute of Molecular Biology, Friedrich-Loeffler-Institut, Südufer 10, 17493 Greifswald-Insel Riems, Germany. Electronic address: axel.karger@fli.bund.de.
authors.forename: Axel
authors.initials: A
authors.lastname: Karger
mesh_terms: D000818:Animals; D001798:Blood Proteins / Q000235:genetics / Q000378:metabolism*; D002417:Cattle / Q000097:blood*; D020543:Proteome; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000662:veterinary*
chemical_list: D001798:Blood Proteins; D020543:Proteome
keywords: Bovine plasma; Combinatorial peptide ligand library; MALDI-TOF/TOF mass spectrometry; Proteomics