Monoclonal antibodies to human neutrophil Fc gamma RIII (CD16) identify polypeptide epitopes.
Human neutrophils constitutively express two low-affinity Fc gamma R, Fc gamma RII (CD32) and Fc gamma RIII (CD16). Eleven monoclonal antibodies (mAb) to CD16 were used to identify antigenic differences among Fc gamma RIII-bearing cells, to define functional epitopes of Fc gamma RIII on neutrophils, and to characterize biochemically the epitopes identified by some of these mAb. Flow cytometry demonstrated that 9 of the 11 mAb reacted with neutrophils, 10 of the 11 reacted with natural killer cells, and 9 of 11 reacted with monocytes and monocyte-derived macrophages. These mAb reacted with CD16 positive cells with varying fluorescence intensities. The ability of anti-CD16 mAb to block the binding of 125I-labeled immune complexes to neutrophils was examined. Four monoclonal antibodies strongly inhibited (87-96%) the binding to neutrophils of 125I-labeled immune complexes. Competitive binding assays were performed to determine whether any other anti-CD16 mAb identify the epitope identified by mAb 3G8. Two other mAb, CLBFCGRAN 1 and CLBGRAN 11, blocked binding of 125I-3G8 IgG to neutrophils. Six of the anti-CD16 mAb efficiently immunoprecipitated polypeptides of broad mobility ranging from 45 to 84 kDa from 125I-labeled neutrophils. When Fc gamma RIII, a complex sialoglycoprotein consisting of almost 50% oligosaccharides, was immunoprecipitated from neutrophils with 3G8 Fab Sepharose and subsequently digested with N-glycanase, 5 of the 6 mAb were capable of immunoprecipitating a deglycosylated polypeptide migrating at 29 kDa. These results demonstrate that these 5 mAb identify polypeptide epitopes of Fc gamma RIII, whereas 1 mAb, YFC120.5, may react with a glycosyl moiety or a determinant whose conformation is dependent on the presence of oligosaccharides.
Fleit
H B
HB
Department of Pathology, State University of New York, Stony Brook 11794-8691.
D000328:Adult; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex / Q000276:immunology; D015703:Antigens, CD / Q000276:immunology*; D000943:Antigens, Differentiation / Q000235:genetics / Q000276:immunology*; D004247:DNA / Q000032:analysis; D000939:Epitopes / Q000032:analysis*; D006801:Humans; D008264:Macrophages / Q000276:immunology; D009000:Monocytes / Q000276:immunology; D009504:Neutrophils / Q000276:immunology*; D011233:Precipitin Tests; D011961:Receptors, Fc / Q000235:genetics / Q000276:immunology*; D017452:Receptors, IgG
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D015703:Antigens, CD; D000943:Antigens, Differentiation; D000939:Epitopes; D011961:Receptors, Fc; D017452:Receptors, IgG; D004247:DNA