Nanopore
Encyclopedia
A nanopore is a small hole. It may, for example, be created by a pore-forming protein or as a hole in synthetic materials such as silicon or graphene.
When a nanopore is present in an electrically insulating membrane
, it can be used as a single-molecule
detector. It can be a biological protein channel in a high electrical resistance lipid bilayer
, a pore in a solid-state membrane or a hybrid of these - 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).
It may be considered a Coulter counter
for much smaller particles.
and MspA porin
. In typical laboratory nanopore experiments, a single protein nanopore is inserted into a lipid bilayer
membrane and single-channel electrophysiology
measurements are taken.
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.
More recently, the use of graphene
as a material for solid-state nanopore sensing has been explored.
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.
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 polymer
s. At this stage, nanopores are making contributions to the understanding of polymer biophysics, as well as to single-molecule analysis of DNA-protein interactions.
.
, glucose
and insulin
to pass however they prevent large immune system molecules like immunoglobins from passing. This way rat pancreatic cells are microencapsulated, they receive nutrients and release insulin through nanopores being totally isolated from their neighboring environment i.e foreign cells. This knowledge can help to replace nonfunctional islets of Langerhans cell
s in the pancreas (responsible for producing insulin), by harvested piglet cells. They can be implanted underneath the human skin without the need of immunosuppressants which put diabetic patients at a risk of infection.
When a nanopore is present in an electrically insulating membrane
Artificial membrane
An artificial membrane, or synthetic membrane, is a synthetically created membrane which is usually intended for separation purposes in laboratory or in industry. Synthetic membranes have been successfully used for small and large-scale industrial processes since the middle of twentieth century. A...
, it can be used as a single-molecule
Molecule
A molecule is an electrically neutral group of at least two atoms held together by covalent chemical bonds. Molecules are distinguished from ions by their electrical charge...
detector. It can be a biological protein channel in a high electrical resistance lipid bilayer
Lipid bilayer
The lipid bilayer is a thin membrane made of two layers of lipid molecules. These membranes are flat sheets that form a continuous barrier around cells. The cell membrane of almost all living organisms and many viruses are made of a lipid bilayer, as are the membranes surrounding the cell nucleus...
, a pore in a solid-state membrane or a hybrid of these - 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
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
) 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).
It may be considered a Coulter counter
Coulter counter
A Coulter counter is an apparatus for counting and sizing particles suspended in electrolytes. It is used for cells, bacteria, prokaryotic cells and virus particles....
for much smaller particles.
Biological/Protein Nanopores
Nanopores may be formed by pore-forming proteins , typically a hollow core passing through a mushroom-shaped protein molecule. Examples of pore-forming proteins are alpha hemolysinHemolysin
Hemolysins are exotoxins produced by bacteria that cause lysis of red blood cells in vitro. Visualization of hemolysis of red blood cells in agar plates facilitates the categorization of some pathogenic bacteria such as Streptococcus and Staphylococcus...
and MspA porin
MspA porin
MspA porin is a membrane porin produced by Mycobacteria, allowing hydrophilic nutrients to enter the bacterium. The protein forms a tightly interconnected octamer with eightfold rotation symmetry that resembles a goblet and contains a central channel. Each subunit fold contains a beta-sandwich of...
. In typical laboratory nanopore experiments, a single protein nanopore is inserted into a lipid bilayer
Lipid bilayer
The lipid bilayer is a thin membrane made of two layers of lipid molecules. These membranes are flat sheets that form a continuous barrier around cells. The cell membrane of almost all living organisms and many viruses are made of a lipid bilayer, as are the membranes surrounding the cell nucleus...
membrane and single-channel electrophysiology
Electrophysiology
Electrophysiology is the study of the electrical properties of biological cells and tissues. It involves measurements of voltage change or electric current on a wide variety of scales from single ion channel proteins to whole organs like the heart...
measurements are taken.
Solid State Nanopores
Solid-state nanopores are generally made in siliconSilicon
Silicon is a chemical element with the symbol Si and atomic number 14. A tetravalent metalloid, it is less reactive than its chemical analog carbon, the nonmetal directly above it in the periodic table, but more reactive than germanium, the metalloid directly below it in the table...
compound membranes, one of the most common being silicon nitride
Silicon nitride
Silicon nitride is a chemical compound of silicon and nitrogen. If powdered silicon is heated between 1300° and 1400°C in an atmosphere of nitrogen, trisilicon tetranitride, Si3N4, is formed. The silicon sample weight increases progressively due to the chemical combination of silicon and nitrogen...
. Solid-state nanopores can be manufactured with several techniques including ion-beam sculpting
Ion-beam sculpting
Ion-Beam scultping is a term used to describe a two-step process to make solid-state nanopores. The term itself was coined by Golovchenko and co-workers at Harvard in the paper "Ion-beam sculpting at nanometer length scales." The term refers to the fact that solid-state nanopores are formed by...
and electron beams.
More recently, the use of graphene
Graphene
Graphene is an allotrope of carbon, whose structure is one-atom-thick planar sheets of sp2-bonded carbon atoms that are densely packed in a honeycomb crystal lattice. The term graphene was coined as a combination of graphite and the suffix -ene by Hanns-Peter Boehm, who described single-layer...
as a material for solid-state nanopore sensing has been explored.
Nanopores may also be used to identify analytes other than DNA. Professor Hagan Bayley
Hagan Bayley
John Hagan Pryce Bayley FRS is a British Professor of Chemical Biology at the University of Oxford. Bayley's research is largely based on the study and engineering of transmembrane pore-forming proteins, as well as interests in chemical signal transduction and biomolecular materials. He is the...
’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.
Nanopore Based Sequencing
The observation that a passing strand of DNA 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
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
sequencing
Sequencing
In genetics and biochemistry, sequencing means to determine the primary structure of an unbranched biopolymer...
, other applications include separation of single stranded and double stranded DNA in solution, and the determination of length of polymer
Polymer
A polymer is a large molecule composed of repeating structural units. These subunits are typically connected by covalent chemical bonds...
s. 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 Nanopores
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 LtdIzon Science
Izon Science Ltd. is a nanotechnology company that has developed instrumentation for nano-scale particle analysis. The company’s main products, qViro and qNano are in use in a wide range of research institutes and universities around the world....
.
Alternate Definition
These can be about 20 nm in a diameter. They are integrated into artificially constructed encapsulated cells of silicon wafers. These pores allow small molecules like oxygenOxygen
Oxygen is the element with atomic number 8 and represented by the symbol O. Its name derives from the Greek roots ὀξύς and -γενής , because at the time of naming, it was mistakenly thought that all acids required oxygen in their composition...
, glucose
Glucose
Glucose is a simple sugar and an important carbohydrate in biology. Cells use it as the primary source of energy and a metabolic intermediate...
and insulin
Insulin
Insulin is a hormone central to regulating carbohydrate and fat metabolism in the body. Insulin causes cells in the liver, muscle, and fat tissue to take up glucose from the blood, storing it as glycogen in the liver and muscle....
to pass however they prevent large immune system molecules like immunoglobins from passing. This way rat pancreatic cells are microencapsulated, they receive nutrients and release insulin through nanopores being totally isolated from their neighboring environment i.e foreign cells. This knowledge can help to replace nonfunctional islets of Langerhans cell
Langerhans cell
Langerhans cells are dendritic cells of the skin and mucosa, and contain large granules called Birbeck granules. They are present in all layers of the epidermis, but are most prominant in the stratum spinosum. They also occur in the papillary dermis, particularly around blood vessels, as well as...
s in the pancreas (responsible for producing insulin), by harvested piglet cells. They can be implanted underneath the human skin without the need of immunosuppressants which put diabetic patients at a risk of infection.