Production, characterization, and application of anti-gossypol polyclonal antibodies

This paper describes the development of antibodies for detection of gossypol in cottonseed. Gossypol was linked to LPH (Limulus polyphemus hemolymph), BSA (bovine serum albumin), amines, or amino acids via Schiff base intermediates for the production of immunogen, solid-phase, and gossypol derivatives. Products were stabilized using NaBH(3)CN. Polyclonal antibodies were produced from rabbits after four immunizations with gossypol-LPH. In ELISA, approximately 0.4 mu caused 50% inhibition of binding (I(50)) to gossypol-BSA solid-phase, when gossypol derivatives were used competitively. This antibody showed higher affinity to lysine-derivatized gossypol, hemigossypol, gossypolone, 6, 6'-dimethoxygossypol, and hexamethoxygossypol compared to underivatized counterparts, and this antibody did not recognize naphthalene or naphthol. The ELISA results for acetone-extractable gossypol in cottonseed products (derivatized with L-(+)-lysine) showed good correlation (r(2) = 0.96) with free gossypol results obtained using the official AOCS method.