Hypophysectomized, terrestrial efts of Diemictylus viridescens respond to treatment with prolactin above an estimated threshold of 0.04 i.u. by migrating to water. The protocol for conducting a prolactin assay using the eft is described and the results of an assay of highly purified prolactin are given. In recent years a minimum dose assay method for the estimation of prolactin has been developed using the terrestrial (red eft) stage of the eastern spotted newt, Diemictylus viridescens, as the test organism. A description of the technique is presented here in order that its protocol be available to those interested in the problem. Following metamorphosis, individuals of Diemictylus enter into the terrestrial eft phase which usually lasts for about three years. The animals then return to water and develop the lateralty compressed (keeled) tail, lateral line system and olive pigmentation of the adult.