A protein fraction with prolactin bioactivities was obtained from the culture medium of human fetal pituitary tissue in long term cultures and termed Pasteels human prolactin (PHP) for convenience. An antiserum was prepared against this human prolactin and used to develop a radioimmunoassay for the material in human plasma. The antigen available (220 µg)was tested against antisera to human growth hormone and to human placental lactogen and in radioimmunoassays for these hormones. The results suggested that the human prolactin fraction contained some immunoreactive human growth hormone (1%). The resulting antibodies in the antihuman prolactin serum could be neutralized by the addition of human growth hormone. The human prolactin also generated antibodies, binding ¹³¹I-labelled human prolactin, which were not neutralized by human growth hormone. This binding was inhibited by plasma from a normal male, 2 normal females and by plasma from a lactating woman after breast feeding. An increase in plasma concentration of the inhibitor, immunologically similar to the human prolactin fraction, was obtained after phenothiazine injection. In the 7 plasma samples tested for human prolactin only 1 contained a detectable concentration of immunoreactive growth hormone. Immunoreactive human placental lactogen (human chorionic somatomammotropin) or any material cross-reacting with anti-HCS serum was undetectable in these plasma samples by a sensitive radioimmunoassay. It is suggested that human plasmas contain detectable amounts of a material immunologically similar to a prolactin fraction isolated from tissue cultures of fetal pituitaries. The fraction and the plasma inhibitor are distinguishable by immunoassay from human growth hormone and from human placental lactogen.