CRISPR-based test for COVID-19 uses smartphone camera

December 07, 2020 // By Rich Pell
CRISPR-based test for COVID-19 uses smartphone camera
Scientists at Gladstone Institutes, University of California, Berkeley and University of California, San Francisco have outlined the technology for a COVID-19 test based on CRISPR gene-editing technology that uses a smartphone camera to provide accurate results in under 30 minutes.

CRISPR (clustered regularly interspaced short palindromic repeats) is a technology that can be used to edit genes and which excels at specifically identifying genetic material. It is seen as offering an alternative testing strategy to polymerase chain reaction (PCR) - the current gold standard of testing.

The new diagnostic test, say the researchers, can not only generate a positive or negative result, it also measures the viral load (or the concentration of SARS-CoV-2, the virus that causes COVID-19) in a given sample.

"It has been an urgent task for the scientific community to not only increase testing, but also to provide new testing options," says Melanie Ott, MD, PhD, director of the Gladstone Institute of Virology and one of the leaders of the study. "The assay [an investigative procedure in laboratory medicine] we developed could provide rapid, low-cost testing to help control the spread of COVID-19."

One of the issues with using the current PCR testing strategy to test for SARS-CoV-2 is that it requires DNA, while the coronavirus is an RNA virus, which means that the viral RNA must first be converted to DNA. In addition, this technique relies on a two-step chemical reaction, including an amplification step to provide enough of the DNA to make it detectable.

As a result, current tests typically need trained users, specialized reagents, and cumbersome lab equipment, which severely limits where testing can occur and causes delays in receiving results. All CRISPR diagnostics to date have also required that the viral RNA be converted to DNA and amplified before it can be detected, adding time and complexity.

In contrast, say the researchers, their novel approach skips all the conversion and amplification steps, using CRISPR to directly detect the viral RNA.

"One reason we're excited about CRISPR-based diagnostics is the potential for quick, accurate results at the point of need," says /Jennifer Doudna, PhD, who is a senior investigator at Gladstone, a professor at UC Berkeley, president of the Innovative Genomics Institute, and an investigator of the Howard Hughes Medical Institute. "This is especially helpful in places with limited access to testing, or when frequent, rapid testing is needed. It could eliminate a lot of the bottlenecks we've seen with COVID-19."

In January 2020, researchers at the lab working on an RNA detection system for HIV decided to shift their focus to COVID-19 when it became clear that the coronavirus was becoming a bigger issue globally and that testing was a potential pitfall.

"We knew the assay we were developing would be a logical fit to help the crisis by allowing rapid testing with minimal resources," says Parinaz Fozouni, a UCSF graduate student and co-first author of a paper on the study. "Instead of the well-known CRISPR protein called Cas9, which recognizes and cleaves DNA, we used Cas13, which cleaves RNA."

In the new test, the Cas13 protein is combined with a reporter molecule that becomes fluorescent when cut, and then mixed with a patient sample from a nasal swab. The sample is placed in a device that attaches to a smartphone.

If the sample contains RNA from SARS-CoV-2, Cas13 will be activated and will cut the reporter molecule, causing the emission of a fluorescent signal. Then, the smartphone camera - essentially converted into a microscope - can detect the fluorescence and report that a swab tested positive for the virus.

"What really makes this test unique is that it uses a one-step reaction to directly test the viral RNA, as opposed to the two-step process in traditional PCR tests," says Ott, who is also a professor in the Department of Medicine at UCSF. "The simpler chemistry, paired with the smartphone camera, cuts down detection time and doesn't require complex lab equipment. It also allows the test to yield quantitative measurements rather than simply a positive or negative result."

The researchers also say that their assay could be adapted to a variety of mobile phones, making the technology easily accessible.

"We chose to use mobile phones as the basis for our detection device since they have intuitive user interfaces and highly sensitive cameras that we can use to detect fluorescence," says UC Berkeley bioengineer Daniel Fletcher, PhD. "Mobile phones are also mass produced and cost effective, demonstrating that specialized lab instruments aren't necessary for this assay."

When the device was tested using patient samples, the researchers confirmed that it could provide a very fast turnaround time of results for samples with clinically relevant viral loads. In fact, they say, the device accurately detected a set of positive samples in under five minutes. For samples with a low viral load, the device required up to 30 minutes to distinguish it from a negative test.

"Recent models of SARS-CoV-2 suggest that frequent testing with a fast turnaround time is what we need to overcome the current pandemic," says Ott. "We hope that with increased testing, we can avoid lockdowns and protect the most vulnerable populations."

Not only does the new CRISPR-based test offer a promising option for rapid testing, say the scientists, but by using a smartphone and avoiding the need for bulky lab equipment, it has the potential to become portable and eventually be made available for point-of-care or even at-home use. And, it could also be expanded to diagnose other respiratory viruses beyond SARS-CoV-2.

In addition, the high sensitivity of smartphone cameras, together with their connectivity, GPS, and data-processing capabilities, have made them attractive tools for diagnosing disease in low-resource regions.

"We hope to develop our test into a device that could instantly upload results into cloud-based systems while maintaining patient privacy, which would be important for contact tracing and epidemiologic studies," says Ott. "This type of smartphone-based diagnostic test could play a crucial role in controlling the current and future pandemics."

For more, see "Amplification-free detection of SARS-CoV-2 with CRISPR-Cas13a and mobile phone microscopy."

Related articles:
CRISPR-powered device promises 'bedside' DNA testing
Portable, reusable COVID-19 sensor tests in 60 seconds
10-second COVID test uses 3D-printed sensor
Coronavirus field test project uses graphene sensing chip
Multiplexed COVID test sensor may enable at-home diagnosis

 


Vous êtes certain ?

Si vous désactivez les cookies, vous ne pouvez plus naviguer sur le site.

Vous allez être rediriger vers Google.