Antibodies with high affinity for spiroperidol--II. Cross reactivity with iodobenzamide and domperidone

Three radioligands, 3H-spiroperidol (3H-SPD), 3H-domperidone (3H-DOMP) and 125I-iodobenzamide (125I-IBZM), were used to investigate the antibody response to two haptens, aminospiroperidol (NH2SPD) and N-aminophenethylspiroperidol (NAPS). Although structurally different, these three radioligands each bind with high affinity to the D2 dopamine receptor. Antibodies with high affinity for 3H-SPD were elicited in rabbits following immunization with the hapten NH2SPD covalently linked to keyhole limpet hemocyanin (KLH). In addition, antibodies in the rabbit anti-NH2SPD antisera bound 125I-IBZM or 3H-DOMP. Rabbit anti-NH2SPD antibodies that bound 125I-IBZM or 3H-DOMP were found to have higher affinity for IBZM or DOMP, respectively, than for SPD. The binding properties of the anti-NH2SPD antibodies that bound 3H-SPD, 125I-IBZM and 3H-DOMP were characterized using a panel of competitive inhibitors and each radioligand appeared to bind to a distinct subpopulation of anti-NH2SPD antibodies. BALB/c mice were immunized with NH2SPD-KLH or NAPS-KLH. A population of antibodies that bound 3H-SPD and a population of antibodies that bound 3H-DOMP were detected. The population of antibodies that bound 3H-DOMP was found to be heteroclitic for DOMP, since DOMP was a more effective competitive inhibitor than SPD. Binding sites for 125I-IBZM were not detected in either the anti-NH2SPD or the anti-NAPS BALB/c antisera. However, two anti-NAPS monoclonal antibodies, N6-24 and N6-29, that bind 3H-SPD with high affinity (Kd = 10(-9) M), were also found to bind IBZM (Ki = 2 x 10(-7) M) and DOMP (Ki = 2 x 10(-6) M). Although anti-NH2SPD and anti-NAPS antibodies were identified that appeared to bind 3H-SPD, 3H-DOMP or 125I-IBZM with high affinity, none of the populations of polyclonal antibodies or monoclonal antibodies bound all three ligands with high affinity.