Specificity and stability of guinea pig anti-progesterone antibodies

Antibodies against progesterone were induced in guinea pigs of both sexes by injection of progesterone-y beta-hemisuccinate conjugated to bovine serum albumin (BSA) in a ratio of 16 moles of steroid per mole of protein. The concentration of antibody binding sites for progesterone of the animals studied ranged from 5 to 20 microM. The expected heterogeneity of binding affinity for progesterone was observed with two major populations apparently predominating. On bound progesterone with an average affinity greater than 2 X 10(9) M-1 and the other showed an average affinity less than or equal to 6 X 10(6) M-1. The antibodies were fond to be stable to extremes of pH and temperature in serum as well as in solutions of ammonium sulfate precipitates. The antibodies were not stable, however, in a more highly purified form. Attempts to obtain active preparations in high yield by purification beyond the ammonium sulfate step were unsuccessful. Competition studies and direct analysis with radiolabeled steroids showed the high-affinity population to be relatively specific for progesterone binding, whereas other steroids were bound according to the polarity rule indicating that the binding forces are predominantly hydrophobic.