Abstract One aspect of the design of biocompatible synthetic surfaces requires an understanding of both the dynamics of protein interaction and the development of techniques to preferentially control protein adsorption. Our studies focus on self-assembled monolayers (SAMs) that are water stable, highly uniform, and a well-controlled surface composition. These SAMs can be used to modify surface properties of metals, polymers, and many other substrates. We report specific effects of surface functional groups on the spreading and other physiological responses of fibroblasts and neuroblastoma cells. Our studies suggest that surfaces with different compositions modulate the conformation of fibronectin (FN) and thus affect the differentiation responses of fibroblasts and neuronal cells in cell type-specific patterns. Using aqueous Fourier transform infrared (FTIR) attenuated total internal reflection (ATR) techniques we directly evaluate the hypothesis that fibronectin conformation differs on various SAMs, thereby altering binding reactions with cell surface receptors. Our results clearly demonstrate a measure of surface-dependent conformational changes for FN which are correlated by independent biological measurements on cell behavior on these surfaces.