We present a generic patterning process by which biomolecules in a passivated background are patterned directly from physiological buffer to microfabricated surfaces without the need for further processing. First, nitrodopamine-mPEG is self-assembled to selectively render TiO2 patterns non-fouling to biomolecule adsorption on hydrophilic and adhesive glass surfaces. After the controlled TiO2 passivation, the biomolecules can be directly adsorbed from solution in a single step creating large scale micropatterned and highly homogeneous arrays of biomolecules with very high pattern definition. We demonstrate the formation of fluid supported lipid bilayers (SLBs) down to the single μm-level limited only by the photolithographic process. Non-specific adsorption of lipid vesicles to the TiO2 background was found to be almost completely suppressed. The SLB patterns can be further selectively functionalized with retained mobility, which we demonstrate through biotin-streptavidin coupling. We envision this single step patterning approach to be very beneficial for membrane-based biosensors and for pattering of cells on a passivated background with complex, sub-cellular geometries; in each application the adherent areas have a tunable mobility of interaction sites controlled by the fluidity of the membrane.