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The preparation and antigenic properties of estriol 16-glucuronide-bovine serum albumin (BSA) and estradiol 17-glucuronide-BSA conjugates in which the hapten molecule is linked to the carrier protein through an (O-carboxymethyl)oxime bridge at the C-6 position on the steroid nucleus, have been described. Antibodies raised against the two immunogens in the rabbit possessed excellent specificity to estriol 16-glucuronide and estradiol 17-glucuronide, respectively, exhibiting no significant cross-reactions with estrogen glucuronides having a substituent in ring A and no cross-reactivities with free estrogens, their sulfates and related steroids.

Using a radiometric assay the effects of estradiol upon the activity of estradiol 2- and 16 alpha-hydroxylases in MCF-7 human breast cancer cells in culture were studied. After 5 days of treatment and 36 h of withdrawal, incubation in the presence of either 2- or 16 alpha-tritiated substrate was carried out. Estradiol (10 nM) significantly increased 16 alpha-hydroxylase activity (21%, P less than 0.01), while no effects on 2-hydroxylase activity was observed. Treatment for 6 weeks caused a major increase in 16 alpha-hydroxylation (65%, P less than 0.001) and a significant reduction in 2-hydroxylation (21%, P less than 0.05). These effects were not observed in estrogen receptor-negative MDA-MB-231 human breast cancer cells. Our observations suggest that these two metabolic pathways in MCF-7 cells are independently regulated and that this regulation is affected via the estrogen receptor.