Previously we found a unique cell surface antigen [Kat1-antigen (Kat1-Ag)] expressed on rat osteoclasts. In the present study, we focused on the expression of this antigen in preosteoclasts, mononuclear precursors of osteoclasts. Immunohistochemical and immunoelectron microscopic observations of the Kat1-Ag expressed in vivo showed the antigen to be present on mononuclear cells having the morphological features of preosteoclasts. The relationship between Kat1-Ag expression and calcitonin receptor (CTR) expression was examined in detail by a double-detection technique for CTR and Kat1-Ag by use of autoradiography and immunocytochemistry, respectively. In a culture system for forming mononuclear preosteoclast-like cells, almost 100% of the mononuclear cells expressing Kat1-Ag also expressed CTR, demonstrating that Kat1-Ag is a reliable immunological marker for identifying preosteoclasts. Interestingly, a significant number of the CTR-positive mononuclear cells did not express the Kat1-Ag. Detection of these cells expressing CTR but not Kat1-Ag strongly suggests the presence of subpopulations in preosteoclasts. We also obtained evidence suggesting that expression of the Kat1-Ag is initiated during the postmitotic stage of the osteoclast progenitors.