(302c) Discrimination of DNA Structure Using Single Protein Channels Supported on Glass Nanopore Membranes

Abstract. Ion channel recordings using transmembrane protein channel and synthetic nanopores are being applied as single-molecule biosensors and for electrical readout of polynucleotide sequences. Alpha-hemolysin (a-HL), a protein toxin isolated from Staphylococcus aureus has been extensively employed for as a biological pore for these purposes. a-HL self-assembles from monomer peptide in artificial lipid bilayers to form a heptamer nano-scale ion channel across the lipid membrane. Ion channel recordings using a-HL are typically performed using a planer bilayer that spans a ~50 um diameter orifice in a polymer-based membrane such as Teflon. The stability of the bilayer in this configuration is generally limited to several hours. We recently reported the fabrication of a single nanopore embedded in a glass or quartz membrane, and a method of preparing a suspended bilayer across the nanopore orifice. The glass nanopore (GNP) membrane is a single conical shaped nanopore embedded in a ~50 um-thick glass membrane. GNP membranes are nearly ideal supports for ion channel recordings due to the greatly improved stability of the lipid bilayer over the nanopore orifice, and electrical properties (low leakage current and low capacitance). Continuous ion channel recordings with the GNP of over 400 hours, and at potential biases of up to ±0.8 V have been demonstrated [1]. The GNP support is suitable for stochastic detection of single molecules and analysis of DNA structure [2]. We will present a description of the GNP membrane preparation and applications of this methodology in biosensing and analysis.

References

(1) White, R.; Ervin, E.; Yang, T.; Chen, X.; Daniel, S.; Cremer, P.; White, H. J. Am. Chem. Soc. 2007, 129, 11766-11775.

(2) Ervin, E.; Kawano, R.; White, R.; White, H. Anal. Chem. 2008, 80, 2069-2076.