The development of an ELISA-assay for semi-quantitative detection of dihydrogriesenin, a sesquiterpene lactone from Geigeria

Abstract

Certain species of Geigeria contain sesquiterpene lactones which cause vomiting disease in sheep. Dihydrogriesenin (DHG), a sesquiterpene lactone from G. aspera, contains an α-methylene function which can spontaneously react with thiol groups on proteins to form a covalent adduct. A specific antiserum against a DHG-protein adduct can be used to determine the fate of DHG in poisoned animals. The preparation of such an antiserum is reported in this paper. DHG was reacted with cysteine and subsequently coupled to serum albumin using the carbodiimide reaction. When rabbits were immunized with one such conjugate (DHG-bovine serum albumin), it was found that the carrier determinants were immunodominant. A DHG-specific anti-serum of sufficient (ELISA) titre could, however, be obtained by alternating serum albumin carriers for DHG in booster immunizations. The ELISA antigen-antibody reaction could be inhibited by prior reaction of the antisera with cysteinyl-DHG in solution.