Breast cancer (BC) is the most commonly diagnosed cancer among American women; however, the development of post-menopausal BC is significantly lower in African Americans as compared to Caucasians. Hormonal stimulation is important in BC development and differences in the conversion of dehydroepiandrosterone (DHEA) into estrogens may be involved in the lower incidence of post-menopausal BC in African American women. DHEA sulfation by sulfotransferase 2B1b (SULT2B1b) is important in regulating the conversion of DHEA into estrogens in tissues. SULT2B1b is localized in both cytosol and nuclei of some tissues including cancerous and associated-normal breast tissue. Immunohistochemical staining was used to evaluate the total expression and subcellular localization of SULT2B1b in African American and Caucasian breast tissues. Cell fractionation, immunoblot analysis and sulfation assays were used to characterize the subcellular expression and activity of SULT2B1b in BC tissues and T-47D breast adenocarcinoma cells. Immunohistochemical analysis of SULT2B1b showed that African Americans had a significantly greater amount of SULT2B1b in epithelial cells of associated-normal breast tissue as compared to Caucasians. Also, more SULT2B1b in African American associated-normal breast epithelial cells was localized in the nuclei than in Caucasians. Equivalent levels of SULT2B1b were detected in breast adenocarcinoma tissues from both African American and Caucasian women. Nuclei isolation and immunoblot analysis of both BC tissue and human T-47D breast adenocarcinoma cells demonstrated that SULT2B1b is present in nuclei and cytoplasm.