Characterization of Murine Monoclonal Antibodies to Human Interferon‐Gamma (IFN‐γ) and Their Application for Sandwich Enzyme‐Linked Immunosorbent Assay (ELISA)

Abstract

The three murine monoclonal antibodies (MAb), D1G2, D9D10, and D13C8, are specific for human interferon-.gamma. (IFN-.gamma.), but not human IFN-.alpha. and IFN-.beta.. They react weakly with heat-treated IFN-.gamma.. The three antibodies recognize different epitopes of the IFN-.gamma. molecule, as evaluated by antibody-binding inhibition experiments. We have used these three monoclonal antibodies to construct a sandwich enzyme-linked immunosorbent assay (ELISA). The best result was obtained when we used D1G2 or D9D10 MAb as a solid-phase immunosorbent and D1G2 or D9D10 MAb as a tracer. When we measured IFN-.gamma. in sera by a combination of D1G2 (a solid-phase) and D1G2 (a tracer), a result similar to the one by a combination of D9D10 (a solid-phase) and D1G2 (a tracer), was obtained. This may suggest that human IFN-.gamma. exists in oligomeric form. Recombinant human IFN-.gamma. expressed in Escherichia coli is detectable at a concentration of 1 ng/ml in this sandwich ELISA. This assay can be employed for the analysis of the structural characteristics of the human IFN-.gamma. molecule as well as measurement of IFN-.gamma. in human sera and tissue culture fluids.