Development of functional tissues often requires spatially controlled growth of cells over 2D surfaces or 3D substrates to maintain their distinct cellular functions; particularly it is essential for culturing anchorage-dependent cells. In this regard, development of new surfaces/substrates with superior surface properties that could control the cell behavior is of great important and extremely necessary for functional tissue engineering as well as to study how the cells spatially recognize and interact with synthetic material systems. Surface patterning is an approach to modify the surface of biomaterials, either chemically or topographically. Both the approaches are well demonstrated in manipulating cell behaviors such as shape, size, orientation, migration, proliferation, and differentiation. In this article, we review various commonly employed methodologies for use in patterning of biomaterial surfaces/substrates and their suitability in controlling cell behaviors.