AbstractBone cells obtained by digestion of fetal mouse or chicken calvaria were tested for their ability to form or resorb bone in vitro. The isolated cells were precultured for 6 days and subsequently cocultured for 11 days with periosteum‐free noninvaded fetal mouse long bone rudiments. Bone formation and resorption during coculture were evaluated by histology and45Ca release from prelabeled bones. The calvarial origin of cells in cocultures was traced by labeling the cells with3H‐thymidine before coculture, followed by autoradiography.Many osteoblasts and osteoclasts as well as fibroblasts developed frommouseperiosteal cells released late in the sequential digestion procedure and previously denoted as “osteoblastlike” (BL). No or few osteoblasts and osteoclasts but many fibroblasts developed from early released cell fractions that have previously been denoted as “osteoclastlike” (CL).Only osteoblasts and fibroblasts but not osteoclasts developed fromchickencalvarial cell fractions. The osteoblasts developed primarily from cell fractions from the inner layer of the periosteum, previously denoted as “osteoblastlike” (OB). Cells obtained from the outer layer of the periosteum (PF) gave rise mainly to fibroblasts.These studies show that osteoblast and osteoclast precursor cells are maintained in monolayer cultures of periosteal cell fractions. However, sequential digestion ofmousecalvaria does not lead to separation of the two types of bone cells. Rather, osteoclast and osteoblast precursors are released jointly, from the periosteal cell layers closest to the bone surface. In thechickencell fractions osteoclast precursors are absent after preculture, resulting in a more homogeneous population of osteoblast and fibroblast but not osteoc