Bacterial infections have been an escalating problem for healthcare facilities, especially those due to multi-drug resistant bacteria. Infections spread in a plethora of ways, particularly through repeated transfers of bacteria among various surfaces. To better understand the process of bacterial transfer, the mechanism of bacteria-surface interaction as well as the affinity of that interaction must be explored. Recent studies suggest that Van der Waals and electrostatic contributions drive the close-proximity interactions between the bacteria and the substrate surface. Understanding the factors that drive bacteria-substrate interactions provides information on bacterial activity such as cell adhesion, death and biofilm formation. To investigate these mechanisms, this research analyzes the viability and affinity associated with the interactions between E. coli and functionalized surfaces via epifluorescence microscopy, optical trapping and surface energy analysis. Epifluorescence microscopy and optical trapping were simultaneously used to monitor cell viability and probe interactions between single E. coli cells and substrate surfaces. Cell death was observed on modified surfaces while unmodified surfaces demonstrated no influence on cell viability. Contact angle measurements were used to quantify the surface energy of E. coli in solution on substrate surfaces. The surface energy of E. coli in solution decreased more rapidly on functionalized substrates than inert surfaces, indicating cell death or biofilm formation.