Antibodies to the Alpha^ and Alpha2-Selective Antagonists Prazosin and Yohimbine as Probes of the Alpha-Adrenergic Binding Sites

Antibodies were raised against a newly synthesized analog (CP57.609) of the o,-selective antagonist prazosin, and against the a.-selective antagonist, yohimbine, by Immunization of rabbits with antigens prepared by covalent linkage of these Uganda to albumin. Competitive Inhibition of [H]prazosln binding to anti-CP57,609 antiserum by a variety of nnlabeled Uganda revealed a spectrum of antibody specificity, with ar selective agents competing more potently than a,-seiectiTe llgands. In contrast, a,-selective ligands competed more potently with the binding of ['H]yofaimbine to the anti-yohlmbine antiserum than a^-selective agents. These respective antisera were subjected to affinity fractionation on a CP57,609or yohimbine-Sepharose 4B resin. Fractions from the CP57,609 resin were eluted successively with phentolamine (10~'M), prazosin (10 M), and guanidine (5M), and from the yohimbine resin with prazosin (10^M), yohimbine (HMM), and guanldlne (SM). The binding profiles of these fractions differed, and in certain fractions the relative order of potency of adrenergic agents was almost identical to that observed with membrane a-adrenergic receptors. Moreover, using these eluted fractions as immunogens, antisera have been obtained which, in the initial bleeds, already possess antiidiotypic activity. These findings therefore suggest that affinity fractionation of antibodies raised against ar and at-selective antagonists may provide useful analogs for the further study of the ligand recognition properties of a-adrenergic receptors. Additionally, it is probable that antiidiotypic antisera will be developed which will recognize the a-adrenergic binding sites. (Hypertension 4 (suppl II): II-183-II-187, 1982)