Goat Antisera to Bovine Parathyroid Hormone: Characterization of Regional Specificity, and Evolution of Titer and Affinity of the 44-68 Region-Specific Antibody Species

Abstract

Antisera against crude native bovine parathyroid hormone (bPTH) were generated in domestic goats. After primary immunization with 60-200 μg of bPTH, booster injections were given at intervals for 4 years. The resulting antisera contained antibodies directed against 3 regions of the PTH molecule, 1-34, 28-48 and 44-68. In each animal the antibodies specific for the 44-68 region were the most sensitive of the 3 types. Their titer and affinity were monitored serially by using ¹²⁵I-labeled bPTH-(41-84) as radioligand. Antigen doses as low as 2 μg produced significant rises in titer, and titers after 3 μg or 20 μg doses were sometimes equal to or higher than after 100-160 μg. The rise in titer after each boost produced an acute increase in affinity. The affinity also seemed to increase slowly during the interval between boosts. After the fifth booster dose these improvements were less marked. This maturation of antibody affinity in one animal led to an antiserum with sufficient sensitivity to hPTH to be useful in the diagnosis of hyperparathyroid states. These observations suggest that when antigenic recognition sites are looked at singly, the evolution of antibodies to PTH may not be as disorderly as had previously been thought. They also suggest an efficient method for obtaining sensitive antisera with a minimal expenditure of antigen.