Membrane-mimetic phospholipid films with or without cholesterol and varying concentrations of one or more synthetic lipopeptide conjugates were formed on an alkylsilane self-assembled monolayer on glass by a process of lipid vesicle fusion. Using a combination of radiochemical titration with I-125-labeled lipopeptides and atomic force microscopy imaging in an aqueous environment, we have characterized the kinetics of multicomponent monolayer formation and film stability as a function of temperature, lipid alkyl chain length, cholesterol content, and the concentration of solution phase bovine serum albumin. These data confirm that multicomponent membrane-mimetic monolayers can be produced in a predictable fashion by vesicle fusion to an alkylated glass substrate. The stability of these systems, however, is compromised when operating above the phase transition temperature of the lipid component, as well as in the presence of albumin.