A digoxin-like immunoreactive substance (DLIS) has been reported in the amniotic fluid. Since radioimmunoassay kits are standardized using serum-based standards, we hypothesized that measurement of DLIS may be an artifact related to the low protein content of amniotic fluid. We analyzed 12 amniotic fluid samples before and after supplementation with lyophilized human serum. The means ± SDs for DLIS (nmol/l) at protein concentrations of 0, 32 and 63 g/l were 1.4 ± 0.16, 0.6 ± 0.09, and 0.4 ± 0.09 nmol/l, respectively. We, therefore, hypothesize that DLIS in amniotic fluid may in part be explained by a technical artifact.