Alterations in peripheral blood T-lymphocyte subsets have been implicated to play a role in the pathophysiology of estrogen deficiency-induced bone loss in postmenopausal women. The aim of this study was to investigate this hypothesis further by flow cytometric analysis of peripheral blood lymphocyte subpopulations together with bone histomorphometry using ovariectomized (OVX) rats as an experimental estrogen deficiency model. 104 female 6-month-old Wistar rats were either OVX or sham-operated (SHAM). Eight rats served as baseline controls. Groups of 8 SHAM and 8 OVX rats were killed 1, 2, 4, 6, 9, and 12 weeks after surgery. Blood was collected prior to sacrifice. In whole blood samples, subpopulations of peripheral lymphocytes were analyzed by flow cytometry using FITC- or PE-labeled monoclonal antibodies against rat CD5, CD4, and CD8. CD4 and CD8 positive T-lymphocytes were determined by a double-labeling technique. Serum samples were analyzed for estradiol and progesterone. The cancellous bone of the distal femoral metaphysis was analyzed by quantitative bone histomorphometry. Ovariectomy caused a statistically significant fall in serum estradiol and progesterone levels from 2 weeks postovariectomy until the end of the trial. Deterioration of cancellous bone structure and osteopenia in the distal femur of OVX rats became evident at 2 and 4 weeks postovariectomy, respectively. Flow cytometric analysis of peripheral blood lymphocytes revealed that, except for a transient increase in CD4 positive T-lymphocytes in OVX rats relative to SHAM animals at 1 week post-surgery, the number of CD5, CD4, or CD8 positive lymphocytes or the mean fluorescence per cell for these antigens in OVX rats remained unchanged throughout the study. Also the CD4+/CD8+ ratio of peripheral blood T-lymphocytes was not influenced by ovariectomy, and none of these parameters were significantly correlated with serum estradiol or progesterone levels. These results suggest that consistent changes in peripheral blood T-lymphocyte subpopulations are not demonstrable in OVX rats by the techniques applied in this study, and do not seem to play a pathogenetic role in the development of estrogen deficiency-induced bone loss in the rat.