title: Double staining technique using a combination of indirect and direct immunofluorescence with monoclonal antibodies.
abstract: A simple method for double staining by immunofluorescence is described. If for double staining using monoclonal antibodies of the same species only one antibody is conjugated with FITC or TRITC, a combination of indirect and direct immunofluorescence is possible. For cell staining the following incubation steps are carried out: Monoclonal antibody I (unlabelled, mouse), anti-mouse immunoglobulin serum FITC- or TRITC-conjugated, normal mouse serum for blocking of free binding sites of the anti-mouse immunoglobulin, and monoclonal antibody II (mouse) which is conjugated with an alternative fluorochrome. The use of this method is demonstrated for investigation of single cell suspensions (performed as a slide test) and of cryostat sections.
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D000954:Antigens, Surface / Q000032:analysis; D001402:B-Lymphocytes / Q000166:cytology; D002462:Cell Membrane / Q000276:immunology; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D005455:Fluorescent Antibody Technique; D006041:Goats / Q000276:immunology; D007106:Immune Sera; D051379:Mice; D012235:Rhodamines; D013194:Staining and Labeling / Q000379:methods*; D013601:T-Lymphocytes / Q000166:cytology; D013861:Thiocyanates; D014966:Xanthenes
chemical_list: D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D005452:Fluoresceins; D007106:Immune Sera; D012235:Rhodamines; D013861:Thiocyanates; D014966:Xanthenes; C009434:tetramethylrhodamine isothiocyanate; D016650:Fluorescein-5-isothiocyanate