Use of Monoclonal Antibodies as Sensitive and Specific Probes for Biologically Active Human γ -interferon

• Published in: Proceedings of the National Academy of Sciences - PNAS 1984-08, Vol.81 (16), p.5219-5222
• Main Authors: Chang, Tse Wen, McKinney, Sam, Liu, Victor, Kung, Patrick C., Vilcek, Jan, Le, Junming
• Format: Article
• Language: English
• Subjects: Animals; Antibodies; Antibodies, Monoclonal; Antigen-Antibody Complex; Antivirals; Bioassay; Cloning, Molecular; DNA, Recombinant; Escherichia coli - genetics; Humans; Immunoradiometric assay; Interferon-gamma - analysis; Interferon-gamma - genetics; Interferon-gamma - immunology; Interferons; Mice; Molecules; Monoclonal antibodies; Radioimmunoassay; Reactivity; Serum albumins; Ungulates
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.81.16.5219

Mouse monoclonal antibodies B1 and B3 are specific for natural and Escherichia coli-derived recombinant human γ -interferon (IFN-γ ). The two antibodies recognize different epitopes of the IFN-γ molecule and do not compete with each other's binding. We have used these two antibodies to construct a solid-phase, sandwich immunoradiometric assay for human IFN-γ . 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 with125I and used as tracer. This assay can be completed in about 4 hr and is capable of detecting IFN-γ levels in human serum or tissue culture fluids as low as 0.1 NIH reference unit/ml. Recombinant human IFN-γ 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-γ , since after exposure to pH 2, 37 degrees C, or 56 degrees 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-γ molecule.