A monoclonal antibody (25.5) has been produced that recognises luminal epithelial cells of the rat mammary gland. This antibody together with monoclonal anti-CALLA antibodies, which react with mammary myoepithelial cells, has been used in biochemical, immunocytochemical and flow cytometric studies. Antibody 25.5 bound to proteins of molecular weight 70K and 25K (K = 10(3) Mr) in both the rat milk fat globule membrane and in single cell suspensions prepared from the virgin adult rat mammary gland. Anti-CALLA antibody (J5), recognised a 93-100K protein in the gland extracts, which co-electrophoresed with the CALLA/CD-10 antigen from NALM-6 acute lymphoblastic leukaemia cell line. Antibody 25.5 bound to the luminal surface of rat mammary epithelial cells at all stages of development from neonatal through to pregnancy, lactation and involution. CALLA immunoreactive staining has previously been shown on basally located presumptive myoepithelial cells at all stages of development. Flow cytometric analyses demonstrated that 25.5 and anti-CALLA antibodies stained independent cell populations in suspensions of single cells prepared from purified epithelial elements from the mammary gland of adult virgin rat.