title: Quantification of influenza virus hemagglutinins in complex mixtures using isotope dilution tandem mass spectrometry.
abstract: Influenza vaccination is the primary method for preventing influenza and its severe complications. Licensed inactivated vaccines for seasonal or pandemic influenza are formulated to contain a preset amount of hemagglutinin (HA), the critical antigen to elicit protection. Current methods to establish the HA concentration of vaccines rely on indirect measurements that are subject to considerable experimental variability. We present a liquid chromatography-tandem mass spectrometry (LC/MS/MS) method for the absolute quantification of viral proteins in a complex mixture. Through use of an isotope dilution approach, HA from viral subtypes H1, H3, H5, and B was determined both directly and rapidly. This method can be applied to purified virus preparations, to monovalent bulk concentrates, or to trivalent inactivated influenza vaccines with improved speed, sensitivity, precision, and accuracy. This LC/MS/MS approach may substantially increase the reliability of methods used to quantitate the amount of antigen in seasonal and pandemic influenza vaccines and reduce the time and effort to deliver influenza vaccines for public health use during the next influenza pandemic.
authors.affiliation: National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA 30341, USA.
authors.forename: Tracie L
authors.initials: TL
authors.lastname: Williams
authors.forename: Leah
authors.initials: L
authors.lastname: Luna
authors.forename: Zhu
authors.initials: Z
authors.lastname: Guo
authors.forename: Nancy J
authors.initials: NJ
authors.lastname: Cox
authors.forename: James L
authors.initials: JL
authors.lastname: Pirkle
authors.forename: Ruben O
authors.initials: RO
authors.lastname: Donis
authors.forename: John R
authors.initials: JR
authors.lastname: Barr
mesh_terms: D000956:Antigens, Viral / Q000032:analysis*; D002853:Chromatography, Liquid / Q000379:methods*; D045424:Complex Mixtures / Q000737:chemistry*; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000032:analysis*; D006801:Humans; D007554:Isotopes; D009975:Orthomyxoviridae / Q000737:chemistry*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000956:Antigens, Viral; D045424:Complex Mixtures; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D007554:Isotopes