Validation of radioimmunoassays to measure prostaglandins F2 alpha and E2 in canine endometrium and plasma.

Equilibrium, double-antibody radioimmunoassay (RIA) procedures were validated to measure concentrations of prostaglandin (PG) F2 alpha, and E2 in canine plasma and endometrium. The procedure for measuring PGF2 alpha or PGE2 used [125I]PGF2 alpha or [3H]PGE2, respectively, as radiolabeled antigens. Specificity was demonstrated by determining the relative potency of selected PG. Relative potencies were calculated by dividing the mass of PGF2 alpha or PGE2 producing 50% inhibition of maximum bound tubes (Bo) by the quantity of test PG producing 50% inhibition in the same assay. Only PGF1 alpha and PGE1 had relative potencies of greater than 1.0% in the PGF2 alpha and PGE2 RIA, respectively. To assess the accuracy of the 2 assay systems, known amounts of authentic hormone were added to plasma, tissue, or 0.05M phosphate-buffered saline solution containing 0.1% gelatin (gel-PBSS). When increasing amounts of PGF2 alpha or PGE2 were added to plasma, tissue, or gel-PBSS, concentrations of PG assayed from extracted or extracted plus chromatographed samples were similar. Additionally, PGF1 alpha or PGE1 was not detected in canine plasma or tissue; therefore, extraction was the only purification needed before analysis of PGF2 alpha or PGE2. The intra-assay precision of 10 assay replicates for a low (70% Bo) and high (27% Bo) sample was 16.0% and 6.7% (coefficient of variation), respectively, for the PGF2 alpha assay. The intra-assay precision of 3 assay replicates for a low (80% Bo) and high (32% Bo) sample was 7.6% and 4.4%, respectively, for the PGE2 assay.(ABSTRACT TRUNCATED AT 250 WORDS)