Studies on the contribution of putative anti-carbohydrate antibodies to the specificity of immunofluorescence staining of murine T cells with chicken anti-mouse (Fab')2 reagents.
Despite a large body of immunochemical data indicating that chicken antibodies specifically recognize immunoglobulin-like structures on mammalian T and B cells, the possibility has been raised that the fluorescence staining of murine T cells with chicken antibodies directed against the (Fab')2 fragment of pooled mouse IgG might be due to carbohydrate moieties shared by heavily glycosylated proteins such as fetuin and mucin. Therefore, the presence of antibody activity against these glycoproteins in chicken antisera to mouse IgG (Fab')2 fragments and to MOPC-41 (murine kappa chains) was tested using a double antibody radioimmune assay. No detectable binding of fetuin or mucin was found in our chicken anti(Fab')2 reagent, whereas activity against (Fab')2 fragments was readily detected. In spite of the lack of anti-mucin or anti-fetuin activity, these glycoproteins were capable, along with fetal calf serum, whole mouse IgG, and mouse (Fab')2 fragments, of inhibiting the binding of anti-mouse (Fab')2 to T cells assayed by double immunofluorescence. However, the effective inhibition by fetuin only occurred when very large amounts of the glycoprotein were used, and titration experiments indicated that fetuin, is on a molar basis, at least 60-fold less efficient and inhibitor of the fluorescence reaction than (Fab')2 and nearly 3000-fold less efficient on a molar carbohydrate basis. We conclude that chicken anti-(Fab')2 and anti-mouse kappa chains stain T cells specifically via an immunoglobulin-like determinant and not by anti-carbohydrate activity.
D000818:Animals; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D002241:Carbohydrates / Q000276:immunology*; D002645:Chickens; D005260:Female; D005455:Fluorescent Antibody Technique; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009077:Mucins / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D000509:alpha-Fetoproteins / Q000276:immunology
D002241:Carbohydrates; D007140:Immunoglobulin Fab Fragments; D009077:Mucins; D000509:alpha-Fetoproteins