A novel radioimmunoassay of 16α-hydroxy-dehydroepiandrosterone and its physiological levels

Abstract

16α-Hydroxy-dehydroepiandrosterone (16α-OH-DHEA) belongs to the products of extensive DHEA metabolism in mammalian tissues. It is a precursor of 16α-hydroxylated estrogens, increased levels of which are associated with autoimmune disorders. A highly specific radioimmunoassay of unconjugated 16α-OH-DHEA was developed and evaluated. Polyclonal rabbit antisera were raised against 3β,16α-dihydroxy-17,19-dione-19- O-(carboxymethyloxime) and 3β,16α-dihydroxy-7,17-dione-7- O-(carboxymethyloxime) BSA conjugates. Two methods were used for preparation of the conjugates. Homologous radioiodinated derivatives with tyrosine methyl ester were prepared as tracers. While antisera to 7-CMO cross-reacted with DHEA as much as by 58%, the cross-reaction of the chosen antiserum prepared via 19-oxogroup by micellar conjugation technique with 16β-OH-DHEA was only 0.13% and with all other structurally related steroids, including DHEA were lower than 0.01%. The detection limit was 0.017 pmol (5.7 pg)/tube, the average intra- and inter-assay coefficients of variation were 8.2 and 11.4%, respectively. Mean recovery of serum spiked with 16α-OH-DHEA varied between 80 and 110%, the results were independent on sample dilution. 16α-OH-DHEA concentrations in 18 randomly selected sera, including 6 samples from patients with thyroid cancer were compared with results obtained by earlier GC–MS method. Physiological levels of 16α-OH-DHEA in 316 sera (184 females and 132 males) analyzed so far varied between 0.0 and 1.86 nmol/l.