ULTRASENSITIVE 2-SITE ASSAYS FOR INHIBIN-A AND ACTIVIN-A USING MONOCLONAL-ANTIBODIES RAISED TO SYNTHETIC PEPTIDES

Monoclonal antibodies produced to synthetic peptides of the alpha and beta-A subunits of inhibin were used to develop the first highly sensitive two-site assays for recombinant human inhibin-A and activin-A. The assay for inhibin-A could detect as little as 5 pg/ml. Activin-A gave a 5.2% cross-reactivity in the inhibin assay. The assay for activin-A could detect 0.1 ng/ml and inhibin-A showed a 5.3% cross-reaction in the activin assay. Determination of inhibins and activins in biological fluids is complicated by the presence of multiple molecular forms and an excess of free alpha-subunits. The present study demonstrates the potential of antibodies raised to synthetic peptides to configure more specific two-site assays for inhibins and activins which could facilitate research on these molecules. The monoclonal antibody used as the labelled antibody in these studies was made by immunizing mice with a peptide corresponding to an internal sequence of the beta-A subunit. It appears likely that similar high affinity anti-peptide monoclonals suitable for use in ultrasensitive two-site assays could be made to other molecules if a rigorous screening of the fusion supernatants on the native molecule was carried out. The use of anti-peptide monoclonal antibodies in the development of ultrasensitive immunoassays may be more widely applicable than commonly realised.