A specific and sensitive homologous RIA for ovine placental lactogen (oPL) has been developed. The assay is specific for oPL in that ovine pituitary PRL (oPRL), GH (oGH), and other pituitary hormones, as well as rat, caprine, bovine, monkey, and human PLs exhibit no cross-reaction in the assay. The lower limit of sensitivity of the assay is 1.0 ng/ml. In uterine vein samples oPL was measurable after day 40 of gestation and in peripheral sera after day 48 (3.5 +/- 1.9 ng/ml). Peripheral plasma oPL concentrations reached peak levels by day 131--141 (649 +/- 205--565 +/- 347 ng/ml), then began to decline approximately 5 days before parturition and disappeared rapidly after delivery. No significant circadian variation in serum oPL concentrations was found during a 4-day period of sampling from day 120--125 of gestation. Fetal serum oPL levels ranged from 24--150 ng/ml throughout pregnancy. oPL was measurable in fetal membranes as early as day 20--30 (0.21 +/- 0.06 microgram/g wet wt), whereas the concentration in maternal caruncles at this time was 0.1 +/- 0.06 microgram/g wet wt. In the placentomes, peak concentrations occurred around day 101--130. Peak concentrations of oPL in allantoic fluid were found between day 35--50, with detectable levels as early as day 18. In amniotic fluid, oPL was measurable between day 40--50. Concentrations of oPL in urine from the pregnant ewe never exceeded 5 ng/ml.