A lipid bilayer with gigaohm resistance was fabricated over a single 800 nm pore in a Si3N4 chip using 50 nm liposomes. The nanopore was prefilled with a polyelectrolyte multilayer (PEM) that triggered the spontaneous fusion of the lipid vesicles. Pore-forming peptide melittin was incorporated in the bilayer, and single channel activities were monitored for a period of 2.5 weeks. The long lifetime of the system enabled the observation of the time-dependent stabilization effect of the melittin open state upon bias application.