Assay methods for 6-keto-prostaglandin F_1α in human urine. Comparison of chromatographic techniques with radioimmunoassay and gas chromatography-negative-ion chemical-ionization mass spectrometry

6-Keto-prostaglandin F_1α (6-keto-PGF_1α) in human urine is considered to be a reflection of renal prostacyclin production. Because of the large amounts of unidentified eicosanoid metabolites in urine that may potentially bind to 6-keto-PGF_1α antisera, most radioimmunoassays include chromatographic purification of urine. A comparison of chromatographic techniques and of antisera to 6-keto-PGF_1α for the assay of human urine is described. Gas chromatography-negative-ion chemical-ionization mass spectrometry (GC-NICI-MS) was used as the reference method. Radioimmunoassays were performed with each of four antisera combined with each of three chromatographic purification systems (silicic acid, Sephadex LH-20, reversed-phase high-performance liquid chromatography). There was marked variability in the results; however, there was at least one chromatographic method for each antiserum that yielded results comparable to GC-NICI-MS. Direct radioimmunoassay of urine without chromatography yielded markedly elevated and variable results for the four antisera. In contrast, the four antisera gave very similar results with direct assay of media from isolated perfused organs. Thus, for the radioimmunoassay of 6-keto-PGF_1α in human urine, each antiserum is sensitive to different contaminants in urine and must be individually matched to a chromatographic purification system.