title: Quantitative analysis of acrylamide labeled serum proteins by LC-MS/MS.
abstract: Isotopic labeling of cysteine residues with acrylamide was previously utilized for relative quantitation of proteins by MALDI-TOF. Here, we explored and compared the application of deuterated and (13)C isotopes of acrylamide for quantitative proteomic analysis using LC-MS/MS and high-resolution FTICR mass spectrometry. The method was applied to human serum samples that were immunodepleted of abundant proteins. Our results show reliable quantitation of proteins across an abundance range that spans 5 orders of magnitude based on ion intensities and known protein concentration in plasma. The use of (13)C isotope of acrylamide had a slightly greater advantage relative to deuterated acrylamide, because of shifts in elution of deuterated acrylamide relative to its corresponding nondeuterated compound by reversed-phase chromatography. Overall, the use of acrylamide for differentially labeling intact proteins in complex mixtures, in combination with LC-MS/MS provides a robust method for quantitative analysis of complex proteomes.
authors.affiliation: Fred Hutchinson Cancer Research Center, 1100 Fairview Ave. N., Seattle, WA 98109, USA. vfaca@fhcrc.org
authors.forename: Vitor
authors.initials: V
authors.lastname: Faca
authors.forename: Marc
authors.initials: M
authors.lastname: Coram
authors.forename: Doug
authors.initials: D
authors.lastname: Phanstiel
authors.forename: Veronika
authors.initials: V
authors.lastname: Glukhova
authors.forename: Qing
authors.initials: Q
authors.lastname: Zhang
authors.forename: Matthew
authors.initials: M
authors.lastname: Fitzgibbon
authors.forename: Martin
authors.initials: M
authors.lastname: McIntosh
authors.forename: Samir
authors.initials: S
authors.lastname: Hanash
mesh_terms: D000178:Acrylamides / Q000737:chemistry*; D000465:Algorithms; D000478:Alkylation; D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000032:analysis* / Q000235:genetics; D002250:Carbon Radioisotopes / Q000737:chemistry; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods; D003545:Cysteine / Q000737:chemistry; D003903:Deuterium / Q000737:chemistry; D006801:Humans; D013058:Mass Spectrometry / Q000295:instrumentation / Q000379:methods; D008969:Molecular Sequence Data
chemical_list: D000178:Acrylamides; D001798:Blood Proteins; D002250:Carbon Radioisotopes; D003903:Deuterium; D003545:Cysteine