Natural tissues have complex structural hierarchy and biological heterogeneity. Tissue engineering utilizes biomaterial scaffolds to mimic various natural properties in order to augment regeneration therapies. Current tissue engineering techniques typically incorporate imprecise scaffold geometries with random cell placement. As a result, little can be known about the specific influences of controlled changes in 3D scaffold properties on cell behavior and therefore making the results less predictive of how cell behaves in a 3D environment. Digital micromirror-assisted projection printing (DMD-PP) technology has become a promising tool to develop user-defined precise 3D microenvironments using complex biological components such as cells and native extracellular matrix (ECM). DMD-PP technology has been used in a variety of applications ranging from tissue engineering to cancer cell migration to neural stem cell culture, with the potential of patterning multiple cells in precise 3D locations. This chapter describes the novel use of DMD-PP technology in developing biomaterial scaffolds with tunable Poisson’s ratio, a fundamental aspect of mechanical property of all biomaterials.