title: Strategy for purification and mass spectrometry identification of SELDI peaks corresponding to low-abundance plasma and serum proteins.
abstract: Analysis by SELDI-TOF-MS of low abundance proteins makes it possible to select peaks as candidate biomarkers. Our aim was to define a purification strategy to optimise identification by MS of peaks detected by SELDI-TOF-MS from plasma or serum, regardless of any treatment by a combinatorial peptide ligand library (CPLL). We describe 2 principal steps in purification. First, choosing the appropriate sample containing the selected peak requires setting up a databank that records all the m/z peaks detected from samples in different conditions. Second, the specific purification process must be chosen: separation was achieved with either chromatographic columns or liquid-phase isoelectric focusing, both combined when appropriate with reverse-phase chromatography. After purification, peaks were separated by gel electrophoresis and the candidate proteins were analyzed by nano-liquid-chromatography-MS/MS. We chose 4m/z peaks (9400, 13,571, 13,800 and 15,557) selected for their differential expression between two conditions, as examples to explain the different strategies of purification, and we successfully identified 3 of them. Despite some limitations, our strategy to purify and identify peaks selected from SELDI-TOF-MS analysis was effective.
authors.affiliation: INSERM, U744, Lille, France.
authors.forename: Marie
authors.initials: M
authors.lastname: Fertin
authors.forename: Justine
authors.initials: J
authors.lastname: Burdese
authors.forename: Olivia
authors.initials: O
authors.lastname: Beseme
authors.forename: Philippe
authors.initials: P
authors.lastname: Amouyel
authors.forename: Christophe
authors.initials: C
authors.lastname: Bauters
authors.forename: Florence
authors.initials: F
authors.lastname: Pinet
mesh_terms: D000465:Algorithms; D000595:Amino Acid Sequence; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D001798:Blood Proteins / Q000032:analysis* / Q000302:isolation & purification* / Q000378:metabolism; D004526:Efficiency; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008954:Models, Biological; D009203:Myocardial Infarction / Q000097:blood / Q000175:diagnosis / Q000378:metabolism; D009994:Osmolar Concentration; D010949:Plasma / Q000737:chemistry* / Q000378:metabolism; D011379:Prognosis; D040901:Proteomics / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D015415:Biomarkers; D001798:Blood Proteins