Abstract
Researchers based at the University of Minnesota Medical School have developed two separate tests using CRISPR-Cas9 gene editing technology that can quickly and accurately diagnose SARSCoV-2 infection.
The first test is a lateral flow antigen test designed for rapid, at home use by non-experts and is designed to differentiate between COVID-19 variants. It can produce results within an hour and works in a similar way to a pregnancy test.
The second test uses fluorescence and is more sensitive and more complex. It can differentiate between SARS-CoV-2, influenza A and B, and respiratory syncytial virus. Although this test needs to be done in a laboratory, it can also be completed within an hour, does not require overly specialized equipment and has the potential to be scaled up for large scale testing.
The team used CRISPR-Cas9 to target the ORF8a section of the SARS-CoV-2 genome—the CDC and WHO currently test for sequences in the N and E genes of the virus. The researchers chose this target, because it differs significantly between different coronaviruses.
The lateral flow test is designed for use in the field and uses gold nanoparticles coated with rabbit antibodies. In order to generate a positive result, both fluorescein isothiocyanate (FITC) and biotin have to be bound on the test strip. If only FITC is bound then the test is active, but not positive.
The second test is a fluorescence-based assay that can be run on a RT-PCR machine that can detect the ORF8a sequence of SARS-CoV-2. This test uses a DNA probe with a fluorophore attached that can bind the target after a 20-minute incubation period during which it is cut by Cas9 causing it to light up.
