A specific, sensitive, and reliable radioimmunoassay for serum-conjugated deoxycholic acid has been developed with antiserum obtained after immunization of rabbits with deoxycholic acid--bovine serum albumin conjugate. The displacement curve of glyco [3H] deoxycholic acid was linear on a logit-log plot from 7.5 to 320 pmol of unlabeled glycodeoxycholic acid. At 50% displacement of bound label, the cross-reactivity of taurodeoxycholic acid was 100%, deoxycholic acid 30%, taurocholic acid 3%, and glycocholic acid 2%. No cross-reactivity was observed with free cholic acid, conjugated or free chenodeoxycholic acids and conjugated or free lithocholic acids. Fasting serum-conjugated deoxycholic acid concentrations in 10 healthy volunteers ranged from 0.18 to 0.92 mumol/1. Over a period of 5 hours following 0.5 g oral cholate administration the serum-conjugated deoxycholic acid concentration did not change in 5 fasting healthy persons, whereas an initial increase of serum cholic acid was observed.