A 10,000-dalton calcium-binding protein (10-kd CaBP) has been described in the placentae and yolk sacs of rats and mice. This protein is similar or identical to vitamin D-dependent intestinal CaBP and these proteins have been implicated in the molecular mechanisms of intestinal calcium absorption and transplacental calcium transport. Using an antiserum to the purified 10-kd rat intestinal CaBP, the localization of CaBP in the 16-17-day mouse yolk sac and placenta was studied immunocytochemically with peroxidase-antiperoxidase labelling and quantified by radial immunodiffusion. A high concentration of immunolabelling was observed in the endodermal cells of the intraplacental yolk sac lining the sinuses of Duval. The columnar endodermal cells lining one side of the endodermal sinuses adjacent to fetal vessels contained most of the immunoreactive label. Quantitation by radial immunodiffusion demonstrated a 5.5-fold higher concentration of CaBP in the portion of the placenta containing most of the intraplacental yolk sac than in the maternal half of the placenta. This demonstration of a 10-kd CaBP within the intraplacental yolk sac suggests this protein functions to facilitate placental calcium transport and is the first study which directly supports the hypothesis of a functional role for the sinuses of Duval in calcium transport.