The in vitro phenotype of bovine articular chondrocytes is described. Chondrocytes plated at high density in roller-bottle and dish cultures were maintained in vitro. The major matrix macromolecules, collagen and proteoglycan, synthesized by these cells were characterized during the course of the culture period. The chondrocytes synthesized mainly Type II collagen, which was found predominantly in the cell-associated matrix. The media contained a mixture of Type II and Type III collagens. Type I collagen was detectable in neither the medium nor the cell-associated matrix. The proteoglycan monomers found in media and cell-associated matrix had the same hydrodynamic sizes as monomers synthesized by cartilage slices or those extracted from adult articular cartilage. The majority of proteoglycans synthesized by the cells were found in high molecular weight aggregates which were readily recovered from the media and were extractable from cell-associated matrix with low ionic strength buffers. The results demonstrate the long-term in vitro phenotypic stability of the bovine articular chondrocytes. The advantages of the in vitro system as a model for studying the effects of external agents, such as drugs and vitamins, are discussed.