title: Application of immuno-mass spectrometry to analysis of a bacterial virulence factor.
abstract: Here we describe a novel antibody-based assay that combines specificity of antibody with precision of mass spectral analysis. The assay is carried out in three steps using a single antigen capture and transfer reagent. The first step of the assay involves antibody immobilization. The second step is antigen capture and washing to remove unbound proteins. The third step involves the analysis of the captured antigens by surface enhanced laser desorption ionization time-of-flight mass spectrometry. The assay is facilitated by the ability of a single nonviable bacterial preparation expressing immunoglobulin-binding proteins that enable antibody immobilization, specific capture of fluid-phase antigen, and direct sample transfer to a protein chip for mass spectral analysis. Proof-of-concept studies using a model Streptococcus pyogenes virulence factor, the secreted cysteine protease SpeB, are presented.
authors.affiliation: Medical College of Ohio, Toledo 43614-5806, USA.
authors.forename: Myrna
authors.initials: M
authors.lastname: Saouda
authors.forename: Terrence
authors.initials: T
authors.lastname: Romer
authors.forename: Michael D P
authors.initials: MD
authors.lastname: Boyle
mesh_terms: D000918:Antibody Specificity; D001426:Bacterial Proteins; D003546:Cysteine Endopeptidases / Q000032:analysis* / Q000276:immunology; D007118:Immunoassay; D013058:Mass Spectrometry / Q000379:methods*; D013297:Streptococcus pyogenes / Q000472:pathogenicity*; D014774:Virulence
chemical_list: D001426:Bacterial Proteins; D003546:Cysteine Endopeptidases; C042477:streptopain