The present study investigates the specificity of anti-endometrial antibodies present in serum of women with endometriosis. A two colour indirect immuno-histochemical method was used, so that the antigenic reactivity of endogenous immunoglobulins was blocked and specific anti-endometrial antibodies were readily distinguishable. Serum was collected from women with endometriosis, before and after 6 months of treatment, and from women without the disease. The reactivity of serum with uterine and ectopic endometrium from women with and without the disease was studied. The frequency of anti-endometrial antibodies in the serum of women with endometriosis was higher (P < 0.001) than in control sera. Most antibodies specifically reacted with glands in ectopic and uterine endometrium. Antibody reactivity was strongest with endometrium from control women, compared with uterine and ectopic endometrium from women with endometriosis (P < 0.01). A proportion of sera containing anti-endometrial antibodies also reacted with vascular endothelium. Binding was strongest to vessels in uterine and ectopic endometrium from women with endometriosis compared to endometrium from women without the disease (P < 0.01). The presence of anti-endometrial antibodies was associated with infertility.