A versatile confocal-based laser scanning lithographic method for controlled, high-fidelity two-dimensional and three-dimensional surface patterning of deformable, solvated, photoactive substrates is presented (see Figure; scale bars represent 200 mu m). Patterned substrates functionalized with gradients of different cell-adhesive ligands can be fabricated, providing a tool for investigating cell-biomaterial interactions and the structure-function relationships of tissues.