Ultrasensitive two-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.
Groome
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School of Biological and Molecular Sciences, Oxford Polytechnic, Headington, U.K.
D028341:Activins; D000911:Antibodies, Monoclonal; D007124:Immunoenzyme Techniques; D007163:Immunosorbent Techniques; D007265:Inhibins / Q000032:analysis*; D010455:Peptides / Q000276:immunology; D011994:Recombinant Proteins / Q000032:analysis
D000911:Antibodies, Monoclonal; D010455:Peptides; D011994:Recombinant Proteins; D028341:Activins; D007265:Inhibins