PRODUCTION AND CHARACTERIZATION OF POLYCLONAL ANTI-S-20499 ANTIBODIES- INFLUENCE OF THE HAPTEN STRUCTURE ON STEREOSPECIFICITY

Immunoassays were studied as an alternative to HPLC methods for the st ereoselective determination of a chiral drug, S 20499, a new anxiolyti c compound that is chemically related to buspirone. The production of highly stereospecific polylclonal antibodies was sought following the construction of appropriately optimized hapten-protein conjugates. Thi s process involved the selection of the structure and the length of th e spacer arm used to couple S 20499 to the carrier protein as well as deciding on the location of the coupling site with respect to the chir al center. Two haptens were prepared: one a derivative resembling the original structure of S 20499, with the effective addition of a carbox ylic acid group, and a second with the effective addition of a butanoi c acid moiety that is supposed to favor stereorecognition. Six stereos pecific polyclonal antisera were obtained in rabbits with two groups o f antibody families defined in terms of specificity. Both approaches g ave high levels of stereospecificity (cross-reactivity towards the opt ical antipode of S 20499 ranged from 4.1% to <0.1%). Although it did n ot decrease the mean apparent affinity constant, the longer spacer imp roved antibody specificity by decreasing cross-reactions towards dealk ylated S 20499 derivatives. Hence, the addition of a four carbon atom bridge should be a valuable tool for increasing antibody stereospecifi city with no drawbacks in terms of specificity and affinity. It was al so shown that long immunization periods appear to have no effect on th e stereospecificity of the antibodies obtained.