The interaction of small phospholipid vesicles with well-characterized surfaces has been studied to assess the effect of the surface free energy of the underlying monolayer on the formation of phospholipid/alkanethiol hybrid bilayer membranes (HBMs). The surface free energy was changed in a systematic manner using single-component alkanethiol monolayers and monolayers; of binary mixtures of thiols. The binary surfaces were prepared on gold by self-assembly from binary solutions of the thiols HS-(CH2)(n)-X (n=11, X=CH3 or OH) in THF. Surface plasmon resonance (SPIR), electrical capacitance, and atomic force microscopy (AFM) measurements were used to characterize the interaction of palmitoyl,oleoyl-phosphatidylcholine (POPC) vesicles with the surfaces. For all surfaces examined, it appears that the polar part of surface energy influences the nature of the POPC assembly that associates with the surface. Comparison of optical, capacitance, and AFM data suggests that vesicles can remain intact or partially intact even at surfaces with a contact angle with water of close to 100degrees. In addition, comparison of the alkanethiols of different chain lengths and the fluorinated compound HS-(CH2)(2)-(CF2)(8)-CF3 that characterize with a low value of the polar part of the surface energy suggests that the quality of the underlying monolayer in terms of number of defects has a significant influence on the packing density of the resulting HBM layer.