There is a significant and growing need across the research and medical communities forlow-cost, high-throughput DNA separation and quantification techniques. The isolation of DNA is a prerequisite for many molecular biology techniques and experiments. Although single-molecule techniques such as Elemans’ STM technique afford extremely high sensitivity, so far such experiments have remained within the confines of academic and research laboratories. The primary reasons for this relate to throughput speed, detection efficiencies, and analysis times. For example, in a conventional solution-based single-molecule detection experiment, one can only detect approximately 10 000 molecules per minute, or one molecule every 6 ms. Although this may sound a lot, consider that a small drop of water (B5 ml) contains B1.67 1023 molecules.
At this speed you would need over 100 trillion years to detect all the water molecules in that single drop. Using a novel nanopore array developed by researchers in the UK, expect to be able to detect up to 1 million molecules simultaneously in the same 6 ms time window (and bringing the timeframe for analyzing the molecules in a single water drop down to some 60 billion years—
about five times the estimated age of the universe).
The above example exaggerates a bit, of course. Compared to a water molecule, which is very small (it consists of only three atoms with an overall diameter of less than 0.3 nm), a DNA molecule is very large. In the real world you would never analyze a drop of DNA but much, much smaller quantities; even then, existing methods are considered to be unacceptably slow. Although it is now several years since the completion of the rough draft of the human genome was announced in 2003, much effort is still focused on identifying genes responsible for specific biological functions or diseases, and determining the DNA sequence bearing this information.
‘‘In recent years, the creation of nanochannels or nanopores in thin membranes has attracted much interest because of the potential to isolate and sense single DNA molecules while they translocate through the highly confined channels,’’ Joshua Edel, a lecturer in micro- and nanotechnology at Imperial College London, explains. ‘‘Nanopores for such applications have already been fabricated, but in all studies to date the detection of translocation events is
performed electrically by measuring the ionic current.
Edel’s group, together with collaborators from Drexel University, presented proof-of-concept studies that describe a novel approach for optically detecting DNA translocation events through an array of solid-state nanopores which allows for ultra-high-throughput parallel detection at the single-molecule level.
‘‘The single-molecule studies performed by Guillaume Chansin, a PhD student within my research group, are very exciting and novel from a technological perspective,’’ says Edel. ‘‘Firstly, this work is the first demonstration of using fluorescence detection to monitor translocation events within a nanopore array.Secondly, this is the first true demonstration of an approach leading towards high through put single-molecule detection confined within nanofluidic structures.’’
This technique functions by electrokinetically driving DNA strands through submicron-sized holes (in this case 300 nm) on an aluminium/silicon nitride membrane. During the translocation process, the molecules are confined to the walls of the nanofluidic channels, allowing 100% detection efficiency.Importantly, the opaque aluminium layer acts as an optical barrier between the illuminated region and the analyte reservoir. In these conditions, high-contrast
imaging of single-molecule events can be performed.
Edel notes that the majority of work to date using nanopores utilizes ionic blockage currents to monitor translocation events. ‘‘Unfortunately, simply measuring blockage currents can only be performed in a single pore,’’ he says.‘‘One of our motivations for using optical detection was to ensure we can probe multiple holes simultaneously, allowing for true high-throughput detection.
Our results indicate that it is possible to obtain high spatial resolution DNA analysis while independently controlling the applied voltage that drives the molecules into the nanopore. A critical feature of the generic approach is the possibility of parallelizing molecular analysis by probing an entire array of nanopores under uniform illumination.’’
According to Edel, there are plenty of potential applications including DNA sequencing, fragment sizing, sieving, separations, and rare event diagnostics. ‘‘For example, using analytical technologies that exist today it is essentially impossible to detect a single DNA strand within a standard blood sample (of a few millilitres) within a reasonable time frame. The technology we describe can potentially allow for such detection to be performed both rapidly and efficiently.’’
It seems that nanopore research is an exciting and growing field. Expect optical detection to play a dominant role in the future of this technology.
Featured scientist: Joshua Edel
Organization: Imperial College London, UK
Relevant publication: Guillaume A. T. Chansin, Rafael Mulero, Jongin
Hong, Min Jun Kim, Andrew J. deMello, Joshua B. Edel: Singlemolecule
spectroscopy using nanoporous membranes. Nano Lett., 7,
2901–2906.
Source
Nano-Society
www.rsc.org/nanoscience
At this speed you would need over 100 trillion years to detect all the water molecules in that single drop. Using a novel nanopore array developed by researchers in the UK, expect to be able to detect up to 1 million molecules simultaneously in the same 6 ms time window (and bringing the timeframe for analyzing the molecules in a single water drop down to some 60 billion years—
about five times the estimated age of the universe).
The above example exaggerates a bit, of course. Compared to a water molecule, which is very small (it consists of only three atoms with an overall diameter of less than 0.3 nm), a DNA molecule is very large. In the real world you would never analyze a drop of DNA but much, much smaller quantities; even then, existing methods are considered to be unacceptably slow. Although it is now several years since the completion of the rough draft of the human genome was announced in 2003, much effort is still focused on identifying genes responsible for specific biological functions or diseases, and determining the DNA sequence bearing this information.
‘‘In recent years, the creation of nanochannels or nanopores in thin membranes has attracted much interest because of the potential to isolate and sense single DNA molecules while they translocate through the highly confined channels,’’ Joshua Edel, a lecturer in micro- and nanotechnology at Imperial College London, explains. ‘‘Nanopores for such applications have already been fabricated, but in all studies to date the detection of translocation events is
performed electrically by measuring the ionic current.
Edel’s group, together with collaborators from Drexel University, presented proof-of-concept studies that describe a novel approach for optically detecting DNA translocation events through an array of solid-state nanopores which allows for ultra-high-throughput parallel detection at the single-molecule level.
‘‘The single-molecule studies performed by Guillaume Chansin, a PhD student within my research group, are very exciting and novel from a technological perspective,’’ says Edel. ‘‘Firstly, this work is the first demonstration of using fluorescence detection to monitor translocation events within a nanopore array.Secondly, this is the first true demonstration of an approach leading towards high through put single-molecule detection confined within nanofluidic structures.’’
This technique functions by electrokinetically driving DNA strands through submicron-sized holes (in this case 300 nm) on an aluminium/silicon nitride membrane. During the translocation process, the molecules are confined to the walls of the nanofluidic channels, allowing 100% detection efficiency.Importantly, the opaque aluminium layer acts as an optical barrier between the illuminated region and the analyte reservoir. In these conditions, high-contrast
imaging of single-molecule events can be performed.
Edel notes that the majority of work to date using nanopores utilizes ionic blockage currents to monitor translocation events. ‘‘Unfortunately, simply measuring blockage currents can only be performed in a single pore,’’ he says.‘‘One of our motivations for using optical detection was to ensure we can probe multiple holes simultaneously, allowing for true high-throughput detection.
Our results indicate that it is possible to obtain high spatial resolution DNA analysis while independently controlling the applied voltage that drives the molecules into the nanopore. A critical feature of the generic approach is the possibility of parallelizing molecular analysis by probing an entire array of nanopores under uniform illumination.’’
According to Edel, there are plenty of potential applications including DNA sequencing, fragment sizing, sieving, separations, and rare event diagnostics. ‘‘For example, using analytical technologies that exist today it is essentially impossible to detect a single DNA strand within a standard blood sample (of a few millilitres) within a reasonable time frame. The technology we describe can potentially allow for such detection to be performed both rapidly and efficiently.’’
It seems that nanopore research is an exciting and growing field. Expect optical detection to play a dominant role in the future of this technology.
Featured scientist: Joshua Edel
Organization: Imperial College London, UK
Relevant publication: Guillaume A. T. Chansin, Rafael Mulero, Jongin
Hong, Min Jun Kim, Andrew J. deMello, Joshua B. Edel: Singlemolecule
spectroscopy using nanoporous membranes. Nano Lett., 7,
2901–2906.
Source
Nano-Society
www.rsc.org/nanoscience
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