A Radioimmunoassay for Arthropod Moulting Hormones, Introducing a Novel Method of Immunogen Coupling

Abstract

Inokosteron-26-oic acid was coupled to thyroglobulin in aqueous pyridine by a water-soluble carbodiimide. After exhaustive dialysis and gel filtration on Sephadex G-25 in the presence of sodium dodecylsulfate, a coupling ratio of 164 haptens per molecule of thyroglobulin was determined. In all 3 animals injected with the conjugate, ecdysone-binding antibodies were detected. After 1 booster injection the antiserum could be diluted 1:5000 (1:4000, or 1:2000) in order to get a 50% binding of [3H]ecdysone. The dissociation constant was calculated as 5.8 .times. 10-10 mol/l. The antiserum had a greater affinity for ecdysone and 22-isoecdysone than for all other ecdysteroids and steroids tested.