Harvard and MIT scientists have developed a low-cost, CRISPR-based diagnostic test that allows users to test themselves for the SARS-CoV-2 virus and its many variants using a saliva sample at home , without the need for any additional equipment. The device, called Minimally Instrumented Sherlock (miSHERLOCK), is easy to use and provides results that can be read and verified on a smartphone app within an hour, the researchers said.
“It successfully differentiates between three different types of SARS-CoV-2 in experiments, and can be rapidly reconfigured to detect additional variants, such as delta,” he said.
The device, described in the journal Science Advances, can be assembled for about US$15 using a 3D printer and commonly available components, and reusing the hardware reduces the cost of the individual assay to US$6 each.
“miSHERLOCK eliminates the need to take patient samples to a centralized testing location and greatly simplifies sample preparation steps, giving patients and doctors a faster, more accurate picture of individual and community health, Which is important during an evolving pandemic,” Helena Day said. Puig is a postdoctoral fellow at Harvard University and the Massachusetts Institute of Technology (MIT).
The researcher used a CRISPR-based technique called “specific high sensitivity enzymatic reporter unlocking” (SHERLOCK).
CRISPR technology is a simple yet powerful tool for genome editing. This allows researchers to easily alter DNA sequences and modify gene function. Sherlock uses an enzyme called Cas12a – the molecular scissor of CRISPR – to cut DNA or RNA at specific locations.
The researchers used SHERLOCK to cut SARS-CoV-2 RNA in a specific region of a gene called a nucleoprotein conserved in several types of the virus. When the molecular scissor successfully binds and cuts the nucleoprotein gene, the single-stranded DNA probe is also truncated, producing a fluorescent signal.
They also created additional SHERLOCK assays designed to target a panel of viral mutations in spike protein sequences that represent the three SARS-CoV-2 genetic variants: alpha, beta and gamma. The spike protein is used by the SARS-CoV-2 virus to infect and enter human cells.
The team used saliva instead of nasopharyngeal swab samples for the test because it is easier for users to collect saliva, and studies have shown that SARS-CoV-2 is detectable in saliva in higher numbers in the days following infection. Is. The researchers tested their diagnostic tool using clinical saliva samples from 27 COVID-19 patients and 21 healthy patients.
They found that miSHERLOCK correctly identified COVID-19-positive patients 96 percent of the time and patients without disease 95 percent of the time. The researchers tested its performance against alpha, beta and gamma SARS-CoV-2 variants by spiking healthy human saliva with full-length synthetic viral RNA with mutations representing each variant.
They found that the device was effective at a range of viral RNA concentrations. “One of the great things about the miSHERLOCK is that it’s completely modular. The device is separate from the assay itself, so you can plug in separate assays for the specific sequence of RNA or DNA that you want to detect. Trying to do that,” said Devora Najjar, a research assistant at the MIT Media Lab.
The team designed their tool with low-resource settings in mind, as the pandemic has brought to light the vast disparities in healthcare that exist in different parts of the world.
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