An immortal cell line, MMSV‐1, has been developed which exhibits many features of the common luminal epithelial cell of the human mammary gland. The cell line was developed by microinjection of SV40 DNA into individual cells in selected colonies in cultures of human milk epithelial cells. Immunohistochemical staining shows that the MMSV‐1 cells express keratins 7,8, 18 and 19 homogeneously in organized filaments which lead into well‐developed desmosomes. They do not express vimentin or keratins found in stratified epithelia or keratin 14 found in basal cells in the mammary gland. The PEM mucin, recognized by the antibody HMFG‐1, is also expressed and appears to be processed normally. Fibronectin is detected but shows the punctate pattern typical of cultured normal milk epithelial cells. MMSV‐1 cells show a reduced requirement for added growth factors, including cyclic AMP‐elevating agents, but do not grow in agar or form tumours in nude mice. Since the transfected cells could be selected on the basis of an extended in vitro life span, antibiotic resistance markers were not introduced and the cells remain sensitive to hygromycin and neomycin.