As the delta variant is causing havoc on unprotected people, the capacity to swiftly detect and track new variations of the virus is critical for public health officials. In recent years, researchers have developed a simple and affordable CRISPR-based diagnostic tool that allows people to test themselves for different strains of the SARS-CoV-2 virus in the comfort of their own homes, using only a sample of their saliva.
It is simple to use and produces results that can be read and verified by a smartphone app attached within 1 hour. In tests, sherlock was able to differentiate between three distinct variations of SARS-CoV-2, and it can be quickly modified to identify other variants such as delta. With the use of a 3D printer and readily accessible components, the gadget can be constructed for less than $15, and by reusing the hardware, the cost of individual tests may be brought down to $6 for each test.
Using A Low-Cost Gadget, Individuals May Test Themselves For Different Strains Of SARS-Cov-2
In her role as an instructor in pediatrics at Boston Children’s Hospital with a focus on infectious illnesses, co-first author Rose Lee has been on the front lines of the COVID-19 epidemic for more than a year, according to the study. According to her, her experiences at the clinic served as inspiration for the idea that would eventually become miSHERLOCK.
Specifically, the scientists resorted to a CRISPR-based technique developed in the lab of Wyss Core Faculty member and lead article author Jim Collins, known as “specific high sensitivity enzymatic reporter unlocking,” for the detection component of their diagnosis (SHERLOCK). Rather than using CRISPR’s “molecular scissors” for snipping DNA or RNA at specific locations, SHERLOCK uses a different type of scissors that cuts other pieces of DNA in surrounding areas. This allows SHERLOCK to be totally engineered with the nucleic acid probe molecules in producing a signal that indicates that the target has been successfully snipped.
To do this, the researchers developed a SHERLOCK reaction that cuts SARS-CoV-2 RNA at particular regions of a gene called Nucleoprotein that has been found to be conserved across various versions of the virus. Molecular scissors, an enzyme called Cas12a, are used to cut single-stranded DNA probes, resulting in the production of a fluorescent signal when the molecular scissors successfully attach to and cut the nucleoprotein gene. Additional SHERLOCK assays were developed to target a panel of different viral mutations in the Spike protein sequences. These reflect 3 SARS-CoV-2 genetic variants, named Alpha, Beta, and Gamma. These assays were developed in collaboration with the National Institutes of Health.
After developing assays that could consistently detect viral RNA within the allowed concentration range for FDA-authorized diagnostic tests, the team concentrated its efforts on overcoming what is probably the most challenging problem in diagnostics: the processing of diagnostic samples.
As opposed to nasopharyngeal swab samples, the researchers chose saliva as their sample collection technique because it is more convenient for users to collect saliva. Studies have shown that SARS-CoV-2 is detectable in saliva for a more extended period following infection (up to 14 days). However, unprocessed saliva offers its own set of difficulties: This substance includes enzymes that destroy a wide range of compounds, resulting in a significant proportion of false positives.
To overcome this difficulty, the researchers devised a unique approach. To begin, scientists added two chemicals called EGTA and DTTto saliva. They heated the sample to 95°C for three minutes, which removed the false-positive signals from untreated saliva and split open any viral particles that may have been present. Afterward, they included a constructed porous membrane that was designed to capture RNA on its surface to ultimately be used in conjunction with the SHERLOCK reaction to create an outcome.