Membrane preparations (100,000 g pellet) of rabbit, baboon and tree shrew (Tupaia belangeri) uteri were studied for binding of [3H]prostaglandin E2 ([3H]PGE2). Unbound [3H]PGE2 was separated by filtration through Whatman GF/F filters. Non-specific binding was determined by the amount of radioactivity associated with the filters in the presence of a 100-fold excess of radioinert PGE2. PGE2 bound to membranes could be displaced by some other prostaglandin (PG) molecules: PGE1, 16,16-dimethyl-PGE2, PGA1 and PGF2 alpha, but not by 6-keto-PGF1 alpha, PGD2 or arachidonic acid. No PGE2 binding was detected using either membrane ghosts from red blood cells or liposomes. Apparent equilibrium of the binding was reached by 60 min. There was no difference in dissociation constant (Kd) values between rabbits of different reproductive stages (mean range +/- S.E.M. was from 4.6 +/- 0.3 to 5.5 +/- 1.0 nM), but pregnant baboons showed a significantly lower value (3.3 +/- 0.4 nM) than did cyclic animals (12.0 +/- 2.0 nM). Binding capacity (Bmax) values, in contrast, were different only between oestrous rabbits and other reproductive stages. The small amounts of Tupaia tissue only permitted estimates of the Kd and Bmax values to be made; these were 3.8 nM and 499 fmol/mg protein for oestrous animals and 5.4 nM and 674 fmol/mg protein for animals on day 7 of pregnancy, assuming only one class of sites. The present results demonstrate the presence of specific binding sites for PGE2 in uteri from several species.