Supported phospholipid bilayers can be formed by established methods such as vesicle fusion and the Langmuir-Blodgett (LB) technique However. challenges remain in regards to creating supported bilayers from various lipid compositions. using various support sui laces, and incorporating membrane proteins Here we report a detergent removal method as an alternative means of supported bilayer formation The process consists of three steps (1) incubation of phospholipid-poly(ethylene glycol) (PEG)-grafted glass with lipid-detergent micelles, (2) detergent removal by washing the surface with vesicles, and (3) incubation with the vesicles to complete lipid adsorption These procedures yielded fluid planar bilayers of zwitterionic lipids Because fluid structures were not obtained by vesicle fusion, the detergent seemed necessary to produce the polymer-supported bilayers While anionic phospholipids inhibited the attachment of fluid bilayers in the absence of calcium ions, supported bilayers with almost full mobility were obtained from lipid mixtures containing 10-20 mol % anionic lipids in the presence of calcium ions The incorporation of the anionic lipids in the bulk-facing leaflet was demonstrated by the binding of dye-labeled annexin V