The in vivo patterns of bak gene expression were determined in human tissues using an immunohistochemical approach. Polyclonal antisera were raised in rabbits against a synthetic peptide corresponding to amino acids 14-36 of the human Bak protein, and were shown to be specific by immunoblot analysis of various human tissues and cell lines. Bak immunoreactivity was detected in a wide variety of cell types and was typically present within the cytosol in a punctuate pattern suggestive of association with intracellular organelles. Consistent with a proapoptotic role for the Bak protein, gradients of Bak protein production were observed in the complex epithelia of the nasopharynx, esophagus, colon, and bladder, with Bak immunointensity being highest in the upper layers and relatively low in the basal portions of these epithelia. Similarly, in the myeloid series of hematopoietic cells, Bak immunoreactivity was strongest in the terminally differentiated granulocytes, with only weak immunostaining occurring in most progenitor cells in the bone marrow. Among the other cell types and tissues with prominent Bak immunostaining were: (a) cardiomyocytes; (b) vascular and visceral smooth muscle cells; (c) basal cells of the prostate glands; (d) myoepithelial cells of the mammary glands; (e) distal convoluted tubules of the kidney; (f) epidermal keratinocytes; (g) enterocytes of the small intestine; (h) Sertoli and Leidig cells of the testes; (i) theca interna cells in the ovary; and (j) adrenal cortex (but not adrenal medulla). Nearly all neurons and glial cells of the central nervous system did not contain immunodetectable Bak protein, whereas sympathetic neurons as well as neurons in dorsal root ganglia and their axons were Bak immunopositive. Most circulating peripheral blood lymphocytes were negative for Bak immunostaining, whereas strong Bak immunoreactivity was found frequently in lymphocytes in the nodes and spleen. Overall, these patterns of bak expression are unique compared to other members of the bcl-2 gene family, and suggest that bak regulates cell death at specific stages of cell differentiation through tissue-specific control of its expression.