A novel parallel-plate bioreactor has been designed to apply a consistent level of fluid flow-induced shear stress to tissue-engineered articular cartilage in order to improve the matrix composition and mechanical properties and more nearly approximate to that of native tissue. Primary bovine articular chondrocytes were seeded into the bioreactor at high densities (1.7 × 106 cell/cm²) without a scaffold and cultured for two weeks under static, no-flow conditions. A mean fluid flow-induced shear stress of 1 dyne/cm² was then applied continuously for 3 days. The application of flow produced constructs with significantly (p < 0.05) higher amounts of total collagen (via hydroxyproline) and specifically type II collagen (via ELISA) (25.3 ± 2.5% and 22.1 ± 4.7% of native tissue, respectively) compared to static controls (22.4 ± 1.7% and 9.5 ± 2.3%, respectively). Concurrently, the tensile Young's modulus and ultimate strength were significantly increased in flow samples (2.28 ± 0.19 MPa and 0.81 ± 0.07 MPa, respectively) compared to static controls (1.55 ± 0.10 MPa and 0.62 ± 0.05 MPa, respectively). This study suggests that flow-induced shear stresses and/or enhanced mass transport associated with the hydrodynamic environment of our novel bioreactor may be an effective functional tissue-engineering strategy for improving matrix composition and mechanical properties in vitro.