The purpose of the present study was to determine whether peripheral blood lymphocytes (PBL) from primary open angle glaucoma (POAG) patients have reduced 3 alpha-hydroxysteroid dehydrogenase (3 alpha-HSD) activity as was previously found in POAG-derived cultured trabecular meshwork cells. The availability of PBL from both POAG and control patients makes this a useful system for studying the association of decreased 3 alpha-HSD activity with POAG. PBL were isolated from the venous blood of 17 POAG patients and 22 non-glaucoma controls and assayed for 3 alpha-HSD activity with tritiated 5 beta-dihydrocortisol as a substrate. The mean 3 alpha-HSD activity +/- S.E.M., expressed in comparable units of specific activity, of the POAG derived PBL was 13.8 +/- 1.3 U as compared to 32.8 +/- 2.0 U for control cells. This reduction (> 50%) was statistically significant (P < 0.001). Quantitative immunoblot analysis of PBL indicated that the POAG and control cells, despite their difference in 3 alpha-HSD activity, had nearly identical amounts of 3 alpha-HSD protein. The molecular weight of PBL 3 alpha-HSD from both groups of patients was 38,000, the same as previously reported for human liver. The results of this study show an association of decreased PBL 3 alpha-HSD activity and POAG which was not related to antiglaucoma therapy. The reduced levels of 3 alpha-HSD activity in the readily obtainable PBL may serve as a marker for POAG or those at risk for developing the disease.