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In this study, the authors describe an ultrasensitive saliva-based COVID-19 assay with a 15-minute sample-to-answer time that does not require RNA isolation or laboratory equipment. This assay uses CRISPR-Cas12a activity to enhance a viral amplicon signal, which is stimulated by the laser diode of a smartphone-based fluorescence microscope device. This device robustly quantified viral load over a broad linear range and exhibited a limit of detection below that of the PCR gold standard assay.
CRISPR data of SARS-CoV-2 RNA levels were similar in saliva and nasal swabs, and viral loads measured by RT-PCR and the smartphone-read CRISPR assay demonstrated good correlation. The development of saliva-based point-of-care diagnostic solutions based on noninvasive procedures can improve access to healthcare and support health monitoring across patients’ lifespans.
– Luiz Meirelles, DDS, MS, PhD
This abstract is available on the publisher's site.
Improved point-of-care COVID-19 assays are urgently needed to improve disease control efforts, but current assays lack the sensitivity of the RT-PCR gold-standard. Herein we describe the development of a portable, ultrasensitive saliva-based COVID-19 assay with a 15 minute sample-to-answer time that does not require RNA isolation or laboratory equipment. This assay uses CRISPR/Cas12a activity to enhance signal from an amplified viral RNA target, which is stimulated by laser diode integrated into a smartphone-based fluorescence microscope readout device. This device robustly quantified viral load over a broad linear range (1-105 copies/μL) and exhibited a limit of detection (0.38 copies/μL) below that of the RT-PCR reference assay. SARS-CoV-2 RNA levels read by CRISPR were similar in patient saliva versus nasal swab samples and viral loads measured by RT-PCR and the smartphone-read CRISPR assay demonstrated good correlation, supporting the potential utility of this portable assay device for saliva-based point-of-care COVID-19 diagnosis.