Biofilm is a polysaccharide matrix embedded with bacteria that forms ubiquitously on almost all kind of surfaces in contact with an aqueous solution. While this biofilm causes a wide range of detrimental effects, including the spread of infectious disease and increased corrosion, the mechanism of biofilm formation at molecular level is poorly understood. In this presentation, we describe the use of surface chemistry (self-assembled monolayers of oligo(ethylene glycol)-terminated alkanethiols on gold) that is known to resist protein adsorption, to control the formation of biofilms. In addition, by fabricating molecular gradients of bio-inertness composed of tri(ethylene glycol) and hydroxyl groups (or methyl groups), we demonstrate that while there is a critical value of surface adhesiveness for mammalian cell adhesion and proliferation, the amount of biofilm formed on surfaces is inversely proportional to surface density of tri(ethylene glycol)-terminated alkanethiols over the entire range of the molecular gradient. This result suggests that biofilm formation is only immobilization-dependent and does not appear to require specific molecular interactions. This immobilization dependence is grossly different from the anchorage dependence of mammalian cell adhesion, which includes specific ligand receptor interaction that triggers a cascade of protein clustering in the cytosol.