<p>To elucidate the mechanisms of cellular mechanotransduction, it is necessary to employ biomaterials that effectively merge biofunctionality with appropriate mechanical characteristics. Agarose is a standard biopolymer used in cartilage mechanobiology but lacks necessary adhesion motifs for cell-matrix interactions to complete mechanostransduction studies. Collagen type I is a natural biomaterial used in cartilage mechanotransduction studies but creates an environment much softer than native cartilage tissue.  In these studies, agarose was blended at two final concentrations (2% w/v and 4% w/v) with collagen type I (2 mg/mL). The overarching goal was to determine whether a composite hydrogel of agarose and collagen can create a mechanically and biologically suitable matrix for chondrocyte studies. First, hydrogels were characterized by rheologic and compressive properties, contraction, and structural homogeneity. Following baseline characterization, primary murine chondrocytes were embedded (1 × 106 cells/mL) within the hydrogels to assess the longer-term <em>in vitro</em> impact on matrix mechanics, cell proliferation, sulfated glycosaminoglycan (sGAG) content, and cellular morphology. To begin probing questions about physiologic loading conditions that chondrocytes experience <em>in</em> <em>vivo</em>, a custom compression loading system was validated using cell-laden hydrogels. Briefly, the 4% agarose – 2 mg/mL collagen I hydrogel composites were able to retain chondrocyte morphology over 21 days in culture, resulted in continual sGAG production, and had bulk mechanics similar to that of the stiffest hydrogel material tested, indicating this hydrogel class may be promising towards developing an effective hydrogel for chondrocyte mechanotransduction and mechanobiology studies, a critical step towards a fuller understanding of cell-matrix interactions. </p>