A biomimetic membrane was obtained by anchoring a hydrophilic monolayer consisting of a triethyleneoxythiol (EO3) or hexaethyleneoxythiol "spacer" to a hanging mercury drop electrode and by interposing a lipid film previously spread on the surface of an aqueous electrolyte between the spacer-coated electrode and the aqueous solution. The impedance spectrum of this biornimetic membrane and its response to the incorporation of the ion carrier valinomycin and of the physiological quinone ubiquinone-10 indicate that it has a good fluidity and consists of a stable lipid bilayer on top of the hydrophilic spacer acting as an ionic reservoir. Upon incorporation of the channel-forming polypeptide melittin, this model system exhibits a conductance entirely analogous to that reported on black lipid membranes, with the appreciable advantage of a higher stability and resistance to disruption. The surface dipole potential of the EO3 monolayer alone was obtained from the extrathermodynamic Tree charge density" q(M) on EO3-coated mercury, as experienced by the diffuse layer ions. The partial charge transfer from the sulfydryl group of EO3 to mercury was estimated from q(M) and from the thermodynamic "total charge density" sigma(M) on EO3-coated mercury.