A team from the University of Washington has developed a device that uses nanotips and electric field manipulation to concentrate and extract DNA from biological samples for subsequent PCR or other analysis.
The group is also working to combine the nanotip technology with fluorescent and electrical detection for specific isolation of target biomarkers for disease diagnostics, starting with a project for rapid point-of-care diagnosis of tuberculosis.
Jae-Hyun Chung, the project's leader and a professor of mechanical engineering at UW, told PCR Insider this week that he set out in his research several years ago to develop a method for concentrating and purifying DNA without the need for centrifugation or microfiltration.
"We identified two main challenges in DNA recovery: one was purification of DNA, and the other was concentration of DNA," Chung said. "Current approaches rely on microfiltration that attracts DNA by change of pH. But when you use those microfilters, DNA is fragmented and lots of information can be lost."
With the group's nanotip device, DNA is concentrated on the nanotip probes via electrical field manipulation in a process that takes only a few minutes. When the nanotip probe is then pulled out of the sample solution, capillary action holds the concentrated DNA molecules onto the structure, Chung explained.
"First, we tested whether the nanotips could concentrate DNA, and they worked really well," he said. "Then, a major concern was whether we could purify DNA in a sample mixture, but we discovered that because the size of the nanotip is very small, when we draw the nanotips out of solution, purification is automatic due to capillary action."
Chung said he began developing the sample-prep device using a UW Center for Commercialization grant of $50,000. Since then, the project has been funded with about $2 million from the National Science Foundation and the National Institutes of Health, according to UW.
The current iteration of the device can work simultaneously on four separate samples, but the researchers believe the technology could be easily scaled to a 96-well plate format.
In research published on the device this year in Analyst, Chung and his colleagues showed that using the nanotip technology to concentrate DNA from saliva samples and then directly dissolving the tip for qPCR yielded comparable DNA concentration results to a commercial DNA extraction kit from Qiagen.
In another study published this year in Analytical and Bioanalytical Chemistry, Chung and his team analyzed the device's ability to concentrate and extract DNA for qPCR analysis from either buccal swabs or saliva samples, again comparing to a Qiagen kit.
According to the researchers, the performance of the nanotip device was equivalent to that of the commercial kit in all the swab samples — with volumes of 5, 10, 50, and 100 μL — and it matched closely in saliva samples as well.
According to Chung, a major advantage of his group's nanotip device is its simplicity and speed.
In the group's Analyst study for example, the nanotip method required only a single step and was able to complete DNA extraction in about six minutes for saliva samples, while the Qiagen kit required 23 steps including centrifugation.
Chung and his colleague Kyong-Hoon Lee have spun out a company, NanoFacture, to commercialize the device. According to UW, Nanofacture is working with KNR Systems in South Korea to manufacture the device appliance while Nanofacture will make the actual nanotips in Washington.
The company is also currently in discussions with potential distribution partners, but Chung said he could not provide additional details. At the same time, the group is developing a simplified, single-tip version of the device that could be used for point-of-care DNA isolation and purification.
Currently, the device concentrates and extracts all DNA in a sample, not particular molecular targets, but Chung said the group is working on nanotips to isolate specific targets for use in molecular diagnostics.
The researchers have developed an approach, described last year in Lab on a Chip, which uses antibody affinity to bind tuberculosis bacteria to a nanotip.
According to Chung, the team has worked initially with fluorescent detection methods for the TB assay, but has also done research indicating that the test could be developed using direct electrical detection.
Chung told PCR Insider that since most target particles are polarized by an electric field, a platform using this methodology should be applicable for various targets including viruses, DNA, proteins, and the like.
In the TB work, Chung and his team have demonstrated that the nanotip method has a detection limit in sputum of 200 M. tuberculosis colony-forming units per mL with a success rate of 96 percent — comparable to PCR.
According to the Lab on a Chip paper, though the device relies on complex physical, chemical, and biological mechanisms, "the simple operation of 'dipping and withdrawal' of tips will allow for screening by minimally trained personnel within 25 minutes. In addition, the minimal power requirement (5 W) combined with low assay cost is ideal for point-of-care screening in limited-resource settings."
The group is now planning to perform a clinical study of the device, Chung said.