A QUANTITATIVE ELISA FOR THE MEASUREMENT OF COMPLEXES OF PROSTATE-SPECIFIC ANTIGEN WITH PROTEIN-C INHIBITOR WHEN USING A PURIFIED STANDARD

The serine protease inhibitor protein C inhibitor is present in semen at a relatively high concentration and forms in vivo complexes with tw o plasminogen activators also present in semen, urokinase-type and tis sue-type plasminogen activators. Therefore, the fact that prostate-spe cific antigen (PSA), a major prostate enzyme, complexes and inactivate s protein C inhibitor (PCI) in semen could have implications in human reproduction. The present study was undertaken to develop an enzyme-li nked immunosorbent assay (ELISA) for complexes of PSA with PCI (PSA:PC I) with purified PSA:PCI complexes as a standard. Seminal plasma was u tilized as the starting material for purification of complexes by affi nity chromatography on heparin-Sepharose and gel filtration. The final preparation contained equimolor concentrations of PSA and PCI and was used for calibration of an ELISA for PSA:PCI complexes involving poly clonal anti-PSA and horseradish peroxidase-labeled anti-PCI antibodies . The ELISA had a detection limit of about 0.2 ng/ml of complex and wa s specific for PSA:PCI complexes because no color was developed at PSA or PCI concentrations up to 100 mug/ml. Normal plasma or plasma from patients with prostate carcinoma who had high PSA levels had no detect able PSA:PCI complexes. Seminal plasma from voluntary donors collected in the absence of inhibitors and incubated at room temperature for at least 3 hours had PSA:PCI complex levels ranging from 30 to 46 mug/ml , accounting for up to 34% of the total PCI in seminal plasma. When se men was collected in the presence of 1,10 phenanthroliniumchloride, to block PSA activity, the mean level of PSA: PCI complex was 11 +/- 6 m ug/ml and the amount of PCI complexed to PSA was 6% +/- 2% of the tota l semen PCI level. The assay therefore makes it possible to carry out further studies on the interaction of PSA and PCI in vivo and in vitro .