In vitro cloning of embryonic cells is a powerful tool for understanding the mechanism and timing of cell differentiation during development. Although in vitro mass culture of chondrogenic mesenchymal cells and clonal culture of already differentiated chondrocytes have been reported, clonal culture of chondrogenic ectomesenchymal cells at a postmigratory stage but prior to chondrogenic cell condensation has not been reported, nor have cartilage nodules been obtained in clonal culture. We cloned chick mandibular ectomesenchymal cells from HH stage 17 chick embryos. These cells produced clones that underwent chondrogenesis as determined by the presence of alcian blue-staining extracellular matrix and immunohistochemical visualization of type II collagen. Chondrogenic clones formed three-dimensional cartilaginous nodules. This postmigratory mandibular ectomesenchyme is heterogeneous, containing two different types of unipotential cells that give rise to chondrogenic and nonchondrogenic cells, plus bipotential cells that give rise to both chondrogenic and nonchondrogenic cells.