Generation and characterization of monoclonal antibodies specific for the human IFN-gamma receptor.
Purified preparations of the human IFN-gamma R derived from placental membranes were used to produce receptor-specific murine mAb. Supernatants from growth-positive wells were screened for their ability to block binding of 125I-IFN-gamma to human placental membranes. Ten inhibitory cultures were identified. Two of these (GIR-208 and GIR-301) abrogated all binding of radioligand to either intact placental membranes or soluble, purified IFN-gamma R. Three others (GIR-72, 76 and 94) showed moderate blocking activity (65, 59, and 49%, respectively) whereas the remaining five (GIR-57, 67, 83, 109, and 153) blocked binding to a low but significant extent (20 to 40%). Specificity experiments demonstrated that the antibodies reacted with the receptor and not the ligand (IFN-gamma). None of the antibodies reacted with IFN-gamma by ELISA. Moreover, GIR-208 and GIR-301, but not isotype-matched controls, identified the receptor by Western blot analysis. GIR-208 and GIR-301 also completely abrogated binding of 125I-IFN-gamma to either mononuclear phagocytes (U937) or human fibroblasts (WISH). Competition experiments revealed that GIR-208 and GIR-301 recognized similar epitopes on the IFN-gamma R and that these (or this) epitopes were identical to or linked to the ligand binding site of the receptor. In addition, both antibodies inhibited development of IFN-gamma-dependent anti-viral activity in WISH cells in a dose-dependent fashion. These data thus indicate that the IFN-gamma R expressed on human placental cells, mononuclear phagocytes, and fibroblasts are similar.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000302:isolation & purification; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D002460:Cell Line; D000939:Epitopes / Q000276:immunology; D005347:Fibroblasts / Q000276:immunology / Q000378:metabolism; D006801:Humans; D007118:Immunoassay; D007371:Interferon-gamma / Q000378:metabolism*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D009000:Monocytes / Q000276:immunology / Q000378:metabolism; D010920:Placenta; D011971:Receptors, Immunologic / Q000276:immunology* / Q000302:isolation & purification; D017471:Receptors, Interferon
D000911:Antibodies, Monoclonal; D000939:Epitopes; D011971:Receptors, Immunologic; D017471:Receptors, Interferon; D007371:Interferon-gamma