Preparation and assessment of antisera to ACTH

Antisera to ACTH were produced in rabbits injected repeatedly at multiple intradermal sites with synthetic [Asp25, Ala26, Gly27]alphah-corticotropin-(1-28)-octacosapeptide-bovine gamma globulin conjugate (octacosapeptide is a sequence analogue of alphah1-28-ACTH). Antibodies to extracted human or porcine ACTH were detected in all of the sera 1 month after immunization. A considerable proportion of the antisera obtained from a single final bleeding 5 months after the primary immunization were suitable for sensitive radioimmunoassay. The antisera were shown to neutralize the steroidogenic activity of ACTH in an isolated rat adrenal cell bioassay system. Titres estimated from antiserum dilution curves and relative avidities from the standard curves were compared. It was possible to detect picogram amounts of ACTH in plasma-free medium with the best antisera. The method described is an effective means of producing anti-sera to the weakly immunogenic N-terminal fragment of the ACTH molecule.