Prolactin receptors present in the particulate fraction of lactating pig mammary gland were solubilized by 7.5mM-3-[(3-cholamidopropyl)dimethylammonio]-1-propane-sulphonic acid (Chaps) and purified by affinity chromatography on prolactin coupled to Affi-Gel 10. Nearly 30% of the particulate receptors were solubilized by the detergent and over a 1000-fold purification from homogenates was achieved. A water-soluble fraction rich in receptors was observed during the preparation of membranes, although this fraction has not yet been purified. Prolactin binding to the receptors was a time-dependent, reversible and saturable reaction in particulate, Chaps-solubilized and purified receptors. In all forms, receptors showed the same specificity to peptide hormones. Prolactin and human growth hormone bound to the same receptors, whereas bovine growth hormone, follicle-stimulating hormone, luteinizing hormone, thyroid-stimulating hormone and insulin failed to bind. After solubilization, the dissociation constant (Kd) for prolactin was decreased 5-fold from 9.8 x 10-11 M in the particulate receptors to 1.8 x 10-'IIM in solubilized and purified receptors, being due principally to an increase in the association rate constant from 1.0 x 109M-1 h-I to (3.9-4.6) x 109M-1 h-', respectively, with the dissociation rate constant remaining unchanged at (1.1-1.3) x 10-2 h-1. Isoelectric focusing of the prolactin-receptor complex revealed two peaks, one at a pl of 5.5-5.6 and the other at 5.2-5.3. Micro-somal receptors were covalently cross-linked to 1251-labelled ovine prolactin with ethylene glycol bis(succinimidyl succinate) and analysed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Autoradiography of the gel revealed a major subunit of M, 28 000-35 000 and a minor one of Mr 67000-69000. Anti-(prolac-tin receptor) antibodies raised against rabbit mammary gland prolactin receptors were equally effective in inhibiting prolactin binding to particulate, solubilized and affinity-purified receptors, suggesting that purified prolactin receptors have a structure indistinguishable immunologically from particulate receptors and rabbit mammary gland prolactin receptors. The present demonstration shows that particulate prolactin receptors from a domestic animal can be solubilized and purified without losing the original properties of high affinity and binding specificity for hormones.