Articular cartilage has multiple histologically distinct longitudinal depth zones. Development and pathogenesis occur throughout these zones. Cartilage explants, monolayer cell culture and reconstituted 3-dimensional cell constructs have been used for investigating mechanisms of pathophysiology in articular cartilage. Such models have been insufficient to reproduce zone-dependent cellular characteristics and extracellular matrix (ECM) upon investigation into cartilage development and pathogenesis. Therefore, we defined a chondrocyte spheroid model consistently formed with isolated chondrocytes from longitudinal depth zones without extrinsic materials. This spheroid showed zone-dependent characteristics of size, cartilage-specific ECM (collagen types I and II, aggrecan and keratan sulfate) and gene expressions of anabolic and catabolic molecules (matrix molecules and matrix metalloproteinase-13). In addition, the spheroid model is small enough to maintain the viability of cells and point symmetry to analyze the gradient of diffusive molecules. This spheroid organoid model will be useful to elucidate the mechanism of histogenesis and pathogenesis in articular cartilage.