title: High confidence and sensitivity four-dimensional fractionation for human plasma proteome analysis.
abstract: Reducing the complexity of plasma proteome through complex multidimensional fractionation protocols is critical for the detection of low abundance proteins that have the potential to be the most specific disease biomarkers. Therefore, we examined a four dimension profiling method, which includes low abundance protein enrichment, tryptic digestion and peptide fractionation by IEF, SCX and RP-LC. The application of peptide pI filtering as an additional criterion for the validation of the identifications allows to minimize the false discovery rate and to optimize the best settings of the protein identification database search engine. This sequential approach allows for the identification of low abundance proteins, such as angiogenin (10(-9) g/L), pigment epithelium growth factor (10(-8) g/L), hepatocyte growth factor activator (10(-7) g/L) and thrombospondin-1 (10(-6) g/L), having concentrations similar to those of many other growth factors and cytokines involved in disease pathophysiology.
authors.affiliation: Department of Medicine, University of Padua, Padua, Italy. millionirenato@gmail.com
authors.forename: Renato
authors.initials: R
authors.lastname: Millioni
authors.forename: Serena
authors.initials: S
authors.lastname: Tolin
authors.forename: Gian Paolo
authors.initials: GP
authors.lastname: Fadini
authors.forename: Marco
authors.initials: M
authors.lastname: Falda
authors.forename: Bas
authors.initials: B
authors.lastname: van Breukelen
authors.forename: Paolo
authors.initials: P
authors.lastname: Tessari
authors.forename: Giorgio
authors.initials: G
authors.lastname: Arrigoni
mesh_terms: D016477:Artifacts; D001798:Blood Proteins / Q000032:analysis*; D005591:Chemical Fractionation / Q000379:methods*; D002852:Chromatography, Ion Exchange; D056148:Chromatography, Reverse-Phase; D030562:Databases, Protein; D005136:Eye Proteins / Q000032:analysis; D006801:Humans; D007525:Isoelectric Focusing; D009414:Nerve Growth Factors / Q000032:analysis; D010455:Peptides / Q000032:analysis; D020543:Proteome / Q000032:analysis*; D012259:Ribonuclease, Pancreatic / Q000032:analysis; D012697:Serine Endopeptidases / Q000032:analysis; D015843:Serpins / Q000032:analysis; D012984:Software; D019700:Thrombospondin 1 / Q000032:analysis; D000099174:Pigment Epithelium-Derived Factor
chemical_list: D001798:Blood Proteins; D005136:Eye Proteins; D009414:Nerve Growth Factors; D010455:Peptides; D020543:Proteome; D012259:Ribonuclease, Pancreatic; D012697:Serine Endopeptidases; D015843:Serpins; D019700:Thrombospondin 1; D000099174:Pigment Epithelium-Derived Factor; C046366:angiogenin; C081092:HGF activator