Cell migration is important for many interesting physiological phenomena including embryonic development, tumor metastasis, and wound healing. The migration behavior of connective tissue cells such as fibroblasts is very difficult to quantify experimentally because these cells migrate relatively slowly. We have developed a technique for creating covalent peptide gradients on surfaces and in biomedically relevant hydrogels to address three challenges: 1. Studying the migration behavior of fibroblasts. 2. Quantifying the activity of specific peptide sequences that may promote migration. 3. Developing platforms for promoting cell migration which can be readily translated to tissue engineering or wound healing applications. We have used these platforms to assay the migration of fibroblasts on surfaces which have been covalently modified to contain gradients of active peptide sequences from the basal lamina protein laminin-1. With this technique, we are able to quantify differences in the migration of fibroblasts: 1. At different absolute peptide concentrations. 2. On peptide gradients with different gradient magnitudes. 3. On peptide gradient surfaces which interact with cells through different adhesion mechanisms. These data provide us with detailed insights into strategies for covalently modifying biomaterials surfaces with peptides in order to maximize cell motility or control the proliferation of a cell type of interest within a tissue engineering scaffold.