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Research Paves The Way For Faster, Better, Cheaper DNA Detection Method

Date:
September 18, 2000
Source:
Northwestern University
Summary:
Researchers at Northwestern University have combined gold nanoparticles and DNA in a novel way to form tiny probes capable of DNA detection that is more accurate and less expensive than conventional detection methods.

EVANSTON, Ill. - Researchers at Northwestern University have combined gold nanoparticles and DNA in a novel way to form tiny probes capable of DNA detection that is more accurate and less expensive than conventional detection methods.

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The new technology, which uses a flatbed scanner to read the results, could displace polymerase chain reaction (PCR) and conventional fluorescence probes in clinical diagnostics and make point-of-care DNA testing possible in the doctor's office and on the battlefield.

Results to be published in the September 8 issue of Science illustrate the method's extraordinary sensitivity and selectivity in the detection of target DNA, in addition to ease of use and low cost.

"Our method, which we call scanometric DNA array detection, may become a truly disruptive technology," said Chad Mirkin, George B. Rathmann Professor of Chemistry and one of the paper's lead authors. "PCR was a disruptive technology when invented in the mid-1980s, and, since then, scientists have been trying to develop DNA detection methods that are as sensitive but without the complexity."

The technology, which Mirkin developed with Robert Letsinger, professor emeritus of chemistry, and Andrew Taton, postdoctoral associate, is 100 times more sensitive than conventional methods, with the ability to detect as few as 60 molecules of DNA without the need for target amplification using methods such as PCR. In addition, the method is so selective it can pinpoint mutations, or bad matches, that would be missed by conventional fluorescence technology.

Once optimized, the technology could be used to quickly and easily detect a wide range of genetic and pathogenic diseases, from cancer and neurodegenerative diseases to HIV and sexually transmitted diseases, as well as biological weapons such as anthrax.

"These tests could be produced for a fraction of what competitor technologies cost," said Mirkin, "and take a fraction of the time to process and interpret results."

The timing of the new technology is perfect, said Mirkin, given the progress being made in genetic sequencing. Understanding the genome will provide vital information for the development of diagnostic and therapeutic technologies related to it, including the Northwestern team's DNA detection method.

Currently, companies in the business of gene chip technology use PCR coupled with fluorescence to do DNA testing. The gene chips are read using a confocal microscope, a complex instrument costing more than $60,000. The whole process requires a long series of complicated steps.

Northwestern's scanometric DNA array detection method could eliminate the expense of PCR and fluorescence, and the results, once amplified using a modified photographic solution, can be read by a flatbed scanner costing as little as $60.

Using this method, various DNA tests can be placed on a glass slide, each test made up of single strands of synthesized DNA, or oligonucleotides, with a sequence designed to bind with its complementary target DNA. The slide then is placed in solution containing the target. At room temperature, perfect and partial matches alike bind to the oligonucleotides on the slide's surface. The gold nanoparticle probes, each covered with 200 oligonucleotide strands, latch on to these pairings.

Next, temperature is used to differentiate the perfect match from a close match. A stringency wash is applied to the slide in which the temperature is raised to just before the melting point of the target DNA. At this point, any mismatches will break apart because partially matched strands are more weakly bound than strands of perfectly matched DNA. Only the perfect matches remain, each with a gold nanoprobe attached and acting as its signal.

The signal is then amplified using modified photographic developing solution. In solution, each gold nanoparticle becomes covered with silver and grows in size, increasing the signal by a factor of 100,000. With current improvements, made with postdoctoral associate Garimella Viswanadham, as few as 60 double-stranded DNA molecules can be detected. A simple flatbed scanner is used to image the slide with a perfect match indicated by a gray dot. The darker the dot, the more target DNA present.

To illustrate the superiority of their method, the researchers took on the challenge of what is called the "G:T wobble," a single-base mismatch of DNA that is difficult for conventional methods to distinguish from the perfect match of the target DNA. Such difficulty in discrimination can lead to false positives.

A comparison of test results showed that the Northwestern method was able to distinguish between the match of two perfectly complementary DNA strands and the near-perfect match where just one base pair was wrong. The fluorescence method was unable to effectively distinguish the two.

"Fluorescence is the workhorse for doing DNA detection, but, with this new method, we now have a superior alternative," said Mirkin. "Our method appears to be better in almost every way ‹ sensitivity, selectivity, cost and convenience."

"We have succeeded in establishing a firm scientific base for the new nanoparticle probe technology," said Letsinger. "We are now collaborating with a company called Nanosphere, Inc. to apply this knowledge in advancing clinical medicine."

In addition to clinical diagnostics, other applications of the scanometric DNA array detection method include pharmacogenomics, forensics and veterinary use.

The research was funded by the U.S. Army Research Office, the National Institutes of Health and Nanosphere, Inc.


Story Source:

The above story is based on materials provided by Northwestern University. Note: Materials may be edited for content and length.


Cite This Page:

Northwestern University. "Research Paves The Way For Faster, Better, Cheaper DNA Detection Method." ScienceDaily. ScienceDaily, 18 September 2000. <www.sciencedaily.com/releases/2000/09/000913212023.htm>.
Northwestern University. (2000, September 18). Research Paves The Way For Faster, Better, Cheaper DNA Detection Method. ScienceDaily. Retrieved October 30, 2014 from www.sciencedaily.com/releases/2000/09/000913212023.htm
Northwestern University. "Research Paves The Way For Faster, Better, Cheaper DNA Detection Method." ScienceDaily. www.sciencedaily.com/releases/2000/09/000913212023.htm (accessed October 30, 2014).

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