‘Glow-In-The-Dark’ Proteins to Help Diagnose Viral Diseases More Quickly and Easily

Despite undergoing technological advancements, most diagnostic tests for viral diseases that are highly sensitive still involve complex techniques to prepare a sample or interpret the results, making it hard to administer these tests in point-of-care settings or in locations with limited resources. Now, a team of researchers has come up with a sensitive technique that analyzes viral nucleic acids in as little as 20 minutes and can be finished in one step using “glow-in-the-dark” proteins.

Bioluminescence is a scientific phenomenon caused by a chemical reaction involving the luciferase protein that creates the luminescent, glow-in-the-dark effect. The luciferase protein has been utilized in creating sensors that emit observable light when they detect their target, making them perfect for point-of-care testing. Nonetheless, these sensors lack the high levels of sensitivity required of a clinical diagnostic test. A gene-editing method known as CRISPR has shown promise in providing this ability, although it requires multiple steps and additional specialized instruments to detect low signals from a complex and noisy sample. So, researchers at Eindhoven University of Technology (Eindhoven, Netherlands) aimed to use CRISPR-related proteins, but combine them with a bioluminescence technique whose signal could be detected with only a digital camera.

In order to ensure that there were sufficient RNA or DNA samples for analysis, the scientists employed recombinase polymerase amplification (RPA), which is a straightforward technique that operates at a constant temperature of around 100 F. The researchers devised a new technique, LUNAS (luminescent nucleic acid sensor), that is comprised of two CRISPR/Cas9 proteins specific for different neighboring parts of a viral genome each have a distinct fragment of luciferase attached to them. Upon detecting the presence of a specific viral genome being tested, the two CRISPR/Cas9 proteins bind to the designated nucleic acid sections and are drawn close to one another, enabling the complete luciferase protein to form and shine blue light in the presence of a chemical substrate.

To take into account the depletion of the substrate, the researchers utilized a control reaction that radiated green light. The presence of a positive result was indicated by a tube that changed from green to blue. The RPA-LUNAS technique successfully detected SARS-CoV-2 RNA in clinical samples obtained from nasal swabs in just 20 minutes, even at concentrations as low as 200 copies per microliter. The researchers believe that the LUNAS assay holds immense potential for quickly and efficiently detecting various other viruses.

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Eindhoven University of Technology