A nanopore is a small hole in an electrically insulating membrane, that can be used as a single-molecule detector. It may be considered a Coulter counter for much smaller particles. It can be a biological protein channel in a high electrical resistance lipid bilayer, a pore in a solid-state membrane or a protein channel set in a synthetic membrane. The detection principle is based on monitoring the ionic current passing through the nanopore as a voltage is applied across the membrane. When the nanopore is of molecular dimensions, passage of molecules (e.g., DNA) cause interruptions of the "open" current level, leading to a "translocation event" signal. The passage of RNA or single-stranded DNA molecules through the membrane-embedded alpha-hemolysin channel (1.5 nm diameter), for example, causes a ~90% blockage of the current (measured at 1 M KCl solution).
Solid-state nanopores are generally made in silicon compound membranes, one of the most common being silicon nitride. Solid-state nanopores can be manufactured with several techniques including ion-beam sculpting and electron beams.
Nanopores may also be used to identify analytes other than DNA. Professor Hagan Bayley’s Research team at the University of Oxford has published research that uses protein nanopores to differentiate between enantiomers of small molecules such as ibuprofen and thalidomide, identify specific biomarkers and screen ion channels. These might have broader applications in clinical medicine and drug development.
The observation that a passing strand of RNA containing different bases results in different blocking levels has led to the nanopore sequencing hypothesis. Oxford Nanopore Technologies and Professor Hagan Bayley's laboratories have shown identification of individual nucleotides including methylated cytosine as they pass through a modified hemolysin nanopore.
Such sequencing, if successful, could revolutionize the field of genomics, as sequencing would be simplified and have the potential for dramatic improvements in power and cost over current versions that use fluorescence/luminescence and optical instrumentation to detect this photon signal. Apart from rapid DNA sequencing, other applications include separation of single stranded and double stranded DNA in solution, and the determination of length of polymers. At this stage, nanopores are making contributions to the understanding of polymer biophysics, as well as to single-molecule analysis of DNA-protein interactions.
Size-tunable elastomeric nanopores have been fabricated, allowing accurate measurement of nanoparticles as they occlude the flow of ionic current.This measurement methodology can be used to measure a wide range of particle types. In contrast to the limitations of solid-state pores, they allow for the optimisation of the resistance pulse magnitude relative to the background current by matching the pore-size closely to the particle-size. As detection occurs on a particle by particle basis, the true average and polydispersity distribution can be determined. Using this principle, the world's only commercial tunable nanopore-based particle detection system has been developed by Izon Science Ltd.
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