Current forays into tissue engineering of articular cartilage in vitro using the self-assembling method have produced constructs possessing significant extracellular matrix and resulting mechanical properties. However, large numbers of native articular chondrocytes are necessary to produce functional engineered cartilage; all previous work with the self-assembling process has used 5.5 x 10(6) cells/construct. In this study, the effects of initial cell seeding (0.25-11 x 10(6) cells/construct) on tissue quality were investigated. Results showed that tissue engineered articular cartilage was formed, when using at least 2 million cells/construct, possessing dimensional, compositional, and compressive properties approaching those of native tissue. It was noted that higher seeding contributed to thicker constructs with larger diameters and had a significant effect on resulting biochemical and biomechanical properties. It was further observed that aggregate modulus increased with increased seeding. By combining gross morphological, histological, biochemical, and biomechanical results, an optimal initial seeding for the self-assembling process of 3.75 x 10(6) cells/construct was identified. This finding enhances the translatability of this tissue engineering process by reducing the number of cells needed for tissue engineering of articular cartilage by 32% while maintaining essential tissue properties.