Osteoblast‐like cells are commonly found in the vicinity of osteoclasts formed in long‐term human bone marrow cultures, and they are believed to be derived from osteogenic cell precursors belonging to the stromal cell system. This paper describes a new culture method for human osteoblasts from the adherent cell population of long‐term human mononuclear bone marrow cultures. The cells obtained exhibited all the classic characteristics of osteoblasts. They contained high intracellular concentrations of alkaline phosphatase and they secreted the osteoblast‐specific marker bone Gla protein. Collagen production was mainly (95–98%) procollagen type I propeptide and only minute quantities of procollagen type III propeptide were detectable by radioimmunoassay in the conditioned medium. After eight weeks the cells formed a mineralized matrix on exposure to β‐glycerophosphate and ascorbic acid. This system provides a model for the study of osteoblast differentiation in vitro and may form the basis for the use of defined media in bone cell cultures due to the presence of high concentrations of osteoblast precursors.