In order to characterize the uterine progesterone-binding proteins of oestrogen-primed and unprimed, ovariectomized immature and adult golden hamsters, cytosols were incubated with [3H] progesterone and/or other steroids and analyzed by sucrose-glycerol density gradient ultracentrifugation. Progesterone-binding components with sedimentation coefficients of 7S and 4.5S were found in the uterine cytosols, but not in the cytosols from the hypothalamus, pituitary, diaphragm, or small intestine. Oestradiol-17β markedly elevated the level of 7S uterine receptor and this increase appeared to be due to new receptor synthesis, since actinomycin D and cycloheximide blocked this response. Fifty to 100 μg of oestradiol-17β per kg body weight was found to promote a maximum increase in the 7S macromolecular component. The 7S receptor showed a tendency to saturate at 1 × 10−7 m [1,2-3H] progesterone, indicating limited capacity. At a molar ratio of 100:1, unlabelled progesterone competed effectively for 7S and 4.5S [3H]progesterone binding, whereas 5α-pregnane-3,20-dione, oestradiol-17β and testosterone did not. Moreover, [1,2-3H] cortisol and [1,2-3H] corticosterone did not bind to the 7S receptor, implying steroid specificity. CI-628, a non-steroid oestrogen antagonist, completely prevented [6,7-3H] oestradiol-17β binding to its 8.5S uterine cytosol receptor, but was entirely without effect on 7S and 4.5S [3H] progesterone binding. Pronase, but not DNase or RNase, abolished 7S and 4.5S progesterone binding, suggesting that the binders are at least in part protein. Protamine sulphate and p-hydroxymercuribenzoate also obliterated 7S binding, implying that this receptor may be an acidic protein which contains sulfhydryl groups that are necessary for progesterone binding.