Study of the removal of 125I-labelled prolactin from the blood of virgin oestrous, pseudopregnant, lactating, weaned and post-partum oestrous rabbits gave a mean half-life of 16·0 ± 1·54 min. No conclusions could be drawn from the results on the influence of physiological condition on half-life. Tissue distribution of 125I-labelled prolactin 60 min after i.v. injection showed that the highest uptake ratio of tissue: plasma was in the kidney, which was confirmed with a concentration of 125I in the urine 6·43 times higher than in the plasma. However, approximately 90% of the 125I in the urine was in the form of labelled products of prolactin degradation. The uptake ratio of mammary tissue: plasma was 2·40 ± 0·09:1 during lactation but was only 0·41 ± 0·04:1 in oestrous rabbits. Of other tissues liver showed the highest ratio of 0·85 ± 0·03:1. The half-life of both 125I- and 131I-labelled prolactin was investigated in vivo in mammary tissue, after i.v. or intraductal injection. No significant differences were seen between methods of administration or differences of isotope, and the average half-life was 52·4 ± 5·9 h. Autoradiographic studies of the tissue localization of 125I- and 131I-labelled prolactin were carried out, the hormone being administered by i.v. or intraductal injection to 15-day pseudopregnant rabbits, or by incubation with 0·5 mm slices of lactating mammary tissue from rabbits. Samples of tissue were obtained from 10 min to 50 h after administration of labelled hormone. Irrespective of method of administration, isotope or time interval, a selective localization of radioactivity was observed on or near to the alveolar secretory cell membrane, on the side adjacent to the vascular supply. It was concluded that specific sites for labelled prolactin occur in this location, since the opposite side of the alveolar cell membrane adjacent to the duct lumen did not show radioactivity localized on or near it, even when the hormone was administered into the teat ducts.