We measured purified extracts of serum (plasma) or urine samples of newborns, pregnant women, normal adults, and uremic patients by a radioreceptor assay (RRA), which uses particulate membrane fractions from human placenta as a binding system, and 125I-digoxin as a tracer. We also measured the digoxin-like immunoreactivity by a sensitive RIA, and results were compared with those found by the RRA. Specific 125I-digoxin binding to placental receptors was competitively inhibited by purified plasma and/or urine extracts of newborns, adult subjects, pregnant women and uremic patients. A linear relationship was found between inhibition of binding and volume of plasma and urine assayed. Moreover, a significant correlation was found between the values obtained by RRA and those found by RIA (n = 17, r = 0.699, p = 0.0012). Our data confirm that increased circulating and/or urinary levels of substances with biological and immunological activity similar to cardiac glycoside drugs are present in newborns, pregnant women and uremic patients compared to healthy adult subjects. In addition, our preliminary study indicates that these endogenous factors are able to bind to the specific receptor of digitalis drugs on the placental membranes.