Use of monoclonal antibodies as sensitive and specific probes for biologically active human gamma-interferon.

Abstract

Mouse monoclonal antibodies B1 and B3 are specific for natural and Escherichia coli-derived recombinant human .gamma.-interferon (IFN-.gamma.). The 2 antibodies recognize different epitopes of the IFN-.gamma. molecule and do not compete with each other''s binding. These 2 antibodies were used to construct a solid-phase, sandwich immunoradiometric assay for human IFN-.gamma.. Purified antibody B1 was coated on polystyrene beads (0.64 cm in diameter) and used as the solid-phase immunoadsorbent, and antibody B3 was labeled with 125I and used as tracer. This assay can be completed in about 4 h and is capable of detecting IFN-.gamma. levels in human serum or tissue culture fluids as low as 0.1 NIH reference unit/ml. Recombinant human IFN-.gamma. derived from E. coli was detectable at a concentration of 0.02 ng/ml. The assay appears to be specific for the biologically active forms of IFN-.gamma., since after exposure to pH 2, 37.degree. C, or 56.degree. C, biological activity and reactivity in the immunoradiometric assay decreased in parallel. The immunoradiometric assay can be employed for the analysis of the structural characteristics of the human IFN-.gamma. molecule.