Monoclonal antibodies directed toward human PRL (hPRL) have been produced by fusion of mouse myeloma cells (Sp2/0-Ag 14) with spleen cells from mice immunized with hPRL. Total immunizing doses of 20 μg and 64 μg hPRL resulted in the production of three highly specific hPRL antibodies. The high affinity antibody, with a Ka value of 0.23 Times 1010 M-1, was used to establish a RIA highly suitable for the measurement of hPRL levels in human serum. The correlation of serum hPRL levels measured using the antibody and those in a conventional rabbit anti-hPRL assay was 0.99 (y = 1.16x - 7.2). These results demonstrate that using the mouse hybridoma technique, it is possible to produce high affinity monospecific monoclonal antibody suitable for the measurement of hPRL in human serum.