Tissue engineering of articular cartilage is a promising alternative to the conventional approaches for cartilage repair. However, recent attempts to develop articular cartilage in vitro have proven to be difficult. The tissue formed in vitro may not accumulate enough extracellular matrix, and the resulting mechanical properties are only a fraction of the native tissue. We investigated whether using specific populations of chondrocytes would improve the properties of the cartilaginous tissue that was generated in vitro. Full-thickness (FT), mid-and-deep zone (MD), and deep-zone (DEEP) chondrocytes were isolated, placed on the surface of porous ceramic substrates and maintained in culture for eight weeks. Tissue developed from DEEP chondrocytes was thicker (FT: 0.94+/-0.03, MD: 0.88+/-0.04, DEEP: 2.4+/-0.1 mm) and had accumulated larger amounts of extracellular matrix (FT: 1.61+/-0.05, MD: 1.5+/-0.1, DEEP: 3.8+/-0.2 mg dry weight) than the tissues formed by the FT and MD chondrocytes. The tissue formed by the FT chondrocytes accumulated the greatest amount of collagen (FT: 211+/-14, MD: 185+/-8, DEEP: 178+/-5 microg/mg dry weight) whereas the tissue formed by the MD chondrocytes accumulated significantly more proteoglycans (FT: 198+/-10, MD: 265+/-10, DEEP: 215+/-5 microg/mg dry weight). Interestingly, MD chondrocytes produced tissue that had compressive mechanical properties up to four times greater than the cartilaginous tissues formed by cells from either the FT or DEEP of cartilage. Thus, a combined population of intermediate and DEEP chondrocytes might be more suitable for the tissue engineering of articular cartilage.