Women with premature ovarian failure (POF) have been reported to have lower serum androgen levels compared with normal women. We reviewed the androgen profiles of 143 POF patients and found androgen levels above normal for postmenopausal women in 16% of these subjects. To determine the source of androgens in those women, we studied the available ovarian biopsy samples of 15 POF patients with increased androgen levels using immunohistochemistry, with a panel of antibodies directed against the main steroidogenic enzymes. Five of the ovarian biopsies exhibited abnormal follicles characterized by hypertrophied theca interna expressing steroidogenic enzymes involved in androgen synthesis. In five other biopsies, the steroidogenic activity was scarce and confined to a small number of ovarian stromal cells, sometimes situated in the proximity of follicular remnants. Finally, in five patients, we found no histological evidence of present or past follicular development beyond the quiescent follicular stage, and no steroidogenic cells were detected by immunohistochemistry. Our findings suggest that ovarian theca-derived cells are a source of androgens in some women with POF, whereas in others, as in most postmenopausal patients, the adrenals or the ovarian hilus cells may synthesize a significant quantity of androgens under LH stimulation.