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 (K_d = 10^-9 M), were also found to bind IBZM (K_i = 2 × 10^-7 M) and DOMP (K_i=2 × 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.