Anti-dihydropyridine Antibodies Exhibit [3H]Nitrendipine Binding Properties Similar to the Membrane Receptor for the 1,4-Dihydropyridine Ca2+ Channel Antagonists

The [3H]nitrendipine binding properties of antidihydropyridine antibodies were characterized and compared with the known [3H]nitrendipine binding properties of the membrane receptor for the 1,4-dihydropyridine Ca2+ channel antagonists. Immunization of rabbits with dihydropyridine-protein conjugates resulted in the production of antibodies with high affinity and specificity for the 1,4-dihydropyridine Ca2+ channel antagonists. Anti-dihydropyridine antibodies were found specifically to bind [3H]nitrendipine, [3H]nimodipine, [3H]nisoldipine, and [3H]PN200–110 with high affinity, while [3H]verapamil, [3H]diltiazem, and [3H]trifluoperazine were not recognized. [3H]Nitrendipine binding to antidihydropyridine antibodies was characterized by a competitive radioimmunoassay. The average dissociation constant of the [3H]nitrendipine-antibody complex was 0.06 ± 0.02 nM for an antiserum studied in detail, and ranged from 0.01 to 0.24 nM for all antisera studied. The average dissociation constant determined from rate constants of association (k1 = 2.7 × 105M−1 s−1) and dissociation (k−1 = 1.8 × 10−4s−1) of [3H]nitrendipine from anti-dihydropyridine antibodies immobilized on Protein-A Sepharose was 0.66 nM. Inhibition of [3H]nitrendipine binding was specific for the 1,4-dihydropyridine Ca2+ channel modifiers, and the concentrations required for half-maximal inhibition ranged between 0.25 and 0.90 nM; structurally unrelated Ca2+ channel antagonists did not modify [3H]nitrendipine binding to the anti-dihydropyridine antibodies. In summary, anti-dihydropyridine antibodies have been shown to have high affinity and specificity for the 1,4-dihydropyridine Ca2+ channel antagonists and to exhibit dihydropyridine binding properties similar to the membrane receptor for the 1,4-dihydropyridine Ca2+ channel antagonists.