title: In vivo protein biotinylation for identification of organ-specific antigens accessible from the vasculature.
abstract: We describe a new methodology, based on terminal perfusion of rodents with a reactive ester derivative of biotin that enables the covalent modification of proteins readily accessible from the bloodstream. Biotinylated proteins from total organ extracts can be purified on streptavidin resin in the presence of strong detergents, digested on the resin and subjected to liquid chromatography-tandem mass spectrometry for identification. In the present study, in vivo biotinylation procedure led to the identification of hundreds of proteins in different mouse organs, including some showing a restricted pattern of expression in certain body tissues. Furthermore, biotinylation of mice with F9 subcutaneous tumors or orthotopic kidney tumors revealed both quantitative and qualitative differences in the recovery of biotinylated proteins, as compared to normal tissues. This technology is applicable to proteomic investigations of the differential expression of accessible proteins in physiological and pathological processes in animal models, and to human surgical specimens using ex vivo perfusion procedures.
authors.affiliation: Swiss Federal Institute of Technology, Zurich, Switzerland.
authors.forename: Jascha-N
authors.initials: JN
authors.lastname: Rybak
authors.forename: Anna
authors.initials: A
authors.lastname: Ettorre
authors.forename: Brigitte
authors.initials: B
authors.lastname: Kaissling
authors.forename: Raffaella
authors.initials: R
authors.lastname: Giavazzi
authors.forename: Dario
authors.initials: D
authors.lastname: Neri
authors.forename: Giuliano
authors.initials: G
authors.lastname: Elia
mesh_terms: D000818:Animals; D000941:Antigens / Q000097:blood*; D014408:Biomarkers, Tumor / Q000097:blood*; D019921:Biotinylation / Q000379:methods*; D002853:Chromatography, Liquid / Q000379:methods; D020869:Gene Expression Profiling / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D009363:Neoplasm Proteins / Q000097:blood*; D009374:Neoplasms, Experimental / Q000097:blood* / Q000276:immunology; D009928:Organ Specificity; D014018:Tissue Distribution
chemical_list: D000941:Antigens; D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins