A simple, rapid and large capacity ELISA for biologically active native and recombinant human IFN gamma

A rapid and sensitive enzyme-immunoassay for native and recombinant human interferon gamma is described. The test is performed in one step at room temperature and is based on the sandwich principle. The IFN gamma preparation is distributed with horse radish peroxidase-labeled monoclonal antibody to IFN gamma in microtiter plates previously coated with a second mab against IFN gamma. The amount of the IFN gamma mab sandwich fixed in the microtiter plate wells is proportional to the color developed after the addition of peroxidase-specific substrate. The two mab's used in the test neutralize IFN gamma and are directed against the same epitope. For this reason they can only detect the biologically active dimeric form of IFN gamma. The IFN gamma-ELISA works in phosphate buffer as well as in tissue culture medium or human serum. As the assay is routinely performed in 2 hours, the limit of detection is 3 U/ml of IFN gamma (0.3 ng/ml). If the assay is performed in 16 hours, the limit of detection decreases to 0.5 U/ml IFN gamma (0.06 ng/ml). The conditions to preserve the activity of IFN gamma preparation as standard are discussed.