We proposed to develop novel, well-defined, synthetic extracellular matrices that support survival and proliferation of human embryonic stem cells. This is an important, unmet need in the field and development of such a substrate would aid in moving stem cell therapies to the clinic. In the second year of support, we continued to make significant progress. Based on our characterization of cell surface receptors for matrix proteins on stem cells completed in the first year, we went on to test specific, purified proteins that will support adhesion and growth of the cells. In addition, we examined whether these substrates will support differentiation of retinal pigmented epithelial cells, an important, clinically relevant cell type that is currently in clinical trials for eye disease. Second, using an interdisciplinary approach, we developed novel hydrogels using click chemistry that support adhesion and proliferation of the stem cells. We continued studies of a cyclic RGD peptide that supports growth of human embryonic stem cells. We then went on to test a new synthetic surface containing another RGD peptide developed by collaborators at Geron and Corning, called synthemax. This surface proved to be efficient at supporting growth of undifferentiated hESC as well as differentiation of useful cell types. We also tested a new liquid version of synthemax. Thus we have identified two synthetic substrates we believe are scalable, inexpensive, and clinically compliant.