Fluorescent immunoprecipitation analysis of cell surface proteins: a methodology compatible with mass-spectrometry.
Radiolabelling and biotinylation of cell proteins followed by immunoprecipitation is a common procedure for biochemical characterization of cell-surface antigens recognized by monoclonal antibodies. Here we present a new method of cell labelling with fluorescent dyes followed by immunoprecipitation and SDS-PAGE with subsequent detection of specific bands by fluorescence imaging devices. Fluorescent immunoprecipitation analysis (FIPA) of cell surface proteins is a fast and sensitive alternative to conventional immunoprecipitation methods, eliminating the need to employ radioactive or biotin labels. The proposed method is compatible with mass spectrometry analysis and permits the identification of immunoprecipitated proteins.
Filatov
Alexander V
AV
Institute of Immunology, Kashirskoye Shosse 24-2, 115478 Moscow, Russia. sorben@online.ru
Krotov
Grigory I
GI
Zgoda
Victor G
VG
Volkov
Yuri
Y
D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid; D005456:Fluorescent Dyes; D006801:Humans; D047468:Immunoprecipitation; D008565:Membrane Proteins / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism*; D008969:Molecular Sequence Data; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry; D020298:U937 Cells
D005456:Fluorescent Dyes; D008565:Membrane Proteins