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A highly specific, quick and sensitive immunenzymometric assay (IEMA) was developed for human interferon-alpha-1 (HuIFN-alpha-1) and human interferon-alpha-2 (HuIFN-alpha-2), respectively. Due to the specificity of the used antibodies it is possible to measure HuIFN-alpha-1 and HuIFN-alpha-2 also in the presence of the corresponding other interferon (IFN) in body fluids as serum or plasma and in culture supernatants. These two-site binding assays are constructed for HuIFN-alpha-1 form tow monoclonal antibodies (ZIM-VD11 and ZIM-IIG11) and for HuIFN-alpha-2 from a sheep antibody against leukocyte-IFN and a monoclonal antibody (ZIM-IIG7). One incubation step is necessary only: for HuLFN-alpha-1, 90 minutes (min), for the determination of HuIFN-alpha-2, 150 min. Very low detection limits characterize these IEMAs: for HuIFN-alpha-1: 0.5 IU/ml, and for HuIFN-alpha-2: 20 IU/ml. Both assays detect the corresponding IFNs as natural and recombinant molecules as well. Only biologically active IFN molecules can be m...