Free-Standing Lipid Bilayers Based on Nanopore Array and Ion Channel Formation.

Integrated nanopores are novel and versatile single-molecule sensors for individual label-free biopolymer detection and characterization. However, their studies and application requires a stable lipid bilayer to maintain protein function. Herein, we describe a method for producing lipid bilayers across a nanopore array on a silicon nitride substrate. We used a painting technique commonly used with Teflon films to embed alpha-hemolysin (alpha-HL) into bilayer lipid membranes (BLMs) to form an ion channel. This was carried out in nanofluid developed in our lab. The membrane formation process, stability of BLMs and ion channel recordings were monitored by patch clamp in real-time. BLM formation was demonstrated by electrical recording (<10 pS conductance) of suspended lipid bilayers spanning a nanopore in the range of +/- 100 mV. Membrane resistance (R-m) and capacitance (C-m) of the device with the bilayer were assessed by membrane test as above 1.0 G Omega and similar to 20 +/- 2 pF, respectively. The silicon nitride surface and aperture edge were smooth at the nanometer lever leading to remarkable membrane stability. The membrane lifetime was 5-24 h. A single alpha-HL channel inserted in 30-60 min applied a potential of +100 mV. The alpha-HL channel currents were recorded at similar to 100 +/- 10 pA. Such integrated nanopores enable analysis of channel functions under various solution conditions from the same BLM. This will open up a variety of applications for ion channels including high-throughput medical screening and diagnosis.