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GE Researchers Use a New PCR Amplification Method to Detect Tuberculosis

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Using different types of amplification enzymes than those currently used in PCR, scientists at GE Global Research say they will be able to reduce the time it takes to diagnose someone with tuberculosis and bring that diagnostic test to the point of care.

Currently, determining whether someone has TB can take up to a month, particularly if that person has a drug-resistant strain, as they grow more slowly in the lab. The process entails getting a sputum sample from a patient, which is then sent to a lab to be grown up for the strain and its resistance status to be determined through cultures and PCR analysis.

"We think we can collapse this entire process to a couple of hours," says Peter Serpentino, the business program manager at GE Healthcare.

To do so, researchers led by John Nelson, a project leader in molecular and cellular biology at GE Global Research, are combining disparate technologies to, as Nelson says, take it "a step beyond PCR."

The first step in the process they are developing is to apply the sputum sample to a paper that is impregnated with chemicals to render the bacterium non-infective and stabilize the DNA. The paper can also be treated so that it changes color once DNA is applied. A punch of that paper is then used in the isothermal amplification process, which uses a mix of enzymes that work at one temperature, 45 degrees Celsius. This mix, Nelson says, follows a strand displacement method in which polymerase creates single-stranded DNA as it shunts along, displacing the previous strand. For this test, they use primers to amplify the hot spots of the tuberculosis genome that confer resistance. "We're interrogating a specific region of the bacterium's genome to see if it is drug resistant," adds Scott Duthie, a molecular biologist at GE.

Once amplified, a color-coded, fluorescence-based reaction can be used to alert the researchers to whether or not the tuberculosis bacterium was present and, if there, to which antibiotics it is resistant. This, Nelson and his team add, could be automated for use at the bedside.

Furthermore, Nelson points out that keeping the temperature stable will allow the test to be used in developing parts of the world.

GE is not the only company interested in better TB diagnostics. In September, researchers from Ce-pheid wrote in a New England Journal of Medicine paper that they had a PCR test in the offing that could detect 98 percent of all cultured-confirmed TB. In addition, the Bill and Melinda Gates Foundation is pouring money — more than $750 million — into TB research.

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