Highly specific antibodies to 13,14-dihydro-15-ketoprostaglandin E2 (PGEM) were raised in rabbits. The animals were immunized with PGEM-bovine serum albumin (BSA)-conjugates. The metabolites were extracted with dichloromethane followed by column chromatography. The final antisera dilution was 1:15000 and the cross-reactivity towards prostaglandin A2, F2 alpha, I2, 13,14-dihydro-15-ketoprostaglandin F2 alpha was less than 0.1%. The limit of detection was 7.8 +/- 4.7 pg/ml plasma at the standard range of 3.9 to 500 pg/ml. The intra- and inter-assay variations were 5 and 12%, respectively. PGEM was measured throughout the menstrual cycle in female volunteers. In normal ovulatory women (n = 6) plasma concentrations of PGEM varied between 0.94 to 2.19 ng/ml. A significant increase of plasma PGEM was detected in the preovulatory phase of the cycle (P less than 0.01) over basal levels. In three of these volunteers cervical mucus was analyzed on PGEM and PGFM concentrations showing a fluctuation from 2 pg to 109 pg for PGEM and 0.05 pg to 2.4 pg for PGFM per ml of cervical mucus. The lowest concentrations have been found at the time of ovulation. The application of the radioimmunological method to the measurement of PGEM in addition to the measurement of prostaglandin E2 may be useful for estimating the turnover rates of this fatty acid.