Cell matrix interactions are important in understanding the healing characteristics of the cornea after refractive surgery or transplantation. The purpose of this study was to characterize in more detail the evolution of biomechanical and optical properties of a stromal equivalent (stromal fibroblasts cultured in a collagen matrix). Human corneal stromal fibroblasts were cultured in a collagen matrix. Compaction and modulus were determined for the stromal equivalent as a function of time in culture and matrix composition. The corneal stromal fibroblasts were stained for α-smooth muscle actin expression as an indicator of myofibroblast phenotype. The nominal modulus of the collagen matrix was 364±41 Pa initial and decreased initially with time in culture and then slowly increased to 177±75 Pa after 21 days. The addition of chondroitin sulfate decreased the contraction of the matrix and enhanced its transparency. Cell phenotype studies showed dynamic changes in the expression of α-smooth muscle actin with time in culture. These results indicate that the contractile behavior of corneal stromal cells can be influenced by both matrix composition and time in culture. Changes in contractile phenotype after completion of the contraction process also indicate that significant cellular changes persist beyond the initial matrix-remodeling phase.