Journal
ADVANCED MATERIALS TECHNOLOGIES
Volume 6, Issue 12, Pages -Publisher
WILEY
DOI: 10.1002/admt.202100602
Keywords
catalase; CRISPR; mobile phone; SARS-CoV-2; viral diagnostics
Categories
Funding
- National Institute of Health [4U54HL119145-08, R01AI118502, R01AI138800, R61AI140489]
- Brigham and Women's Hospital through Innovation Evergreen Fund
- CAPES/Harvard Junior Visiting Professor/Researcher Program
- CNPq-Brazil
- CAPES/Print Program
- Fapesb
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This study introduces an amplification-free CRISPR/Cas12a-based technology for SARS-CoV-2 RNA detection, with results readout using a smartphone camera. The method achieves high analytical sensitivity and accuracy, with results obtained in approximately 71 minutes.
CRISPR (Clustered regularly interspaced short palindromic repeats)-based diagnostic technologies have emerged as a promising alternative to accelerate delivery of SARS-CoV-2 molecular detection at the point of need. However, efficient translation of CRISPR-diagnostic technologies to field application is still hampered by dependence on target amplification and by reliance on fluorescence-based results readout. Herein, an amplification-free CRISPR/Cas12a-based diagnostic technology for SARS-CoV-2 RNA detection is presented using a smartphone camera for results readout. This method, termed Cellphone-based amplification-free system with CRISPR/CAS-dependent enzymatic (CASCADE) assay, relies on mobile phone imaging of a catalase-generated gas bubble signal within a microfluidic channel and does not require any external hardware optical attachments. Upon specific detection of a SARS-CoV-2 reverse-transcribed DNA/RNA heteroduplex target (orf1ab) by the ribonucleoprotein complex, the transcleavage collateral activity of the Cas12a protein on a Catalase:ssDNA probe triggers the bubble signal on the system. High analytical sensitivity in signal detection without previous target amplification (down to 50 copies mu L-1) is observed in spiked samples, in approximate to 71 min from sample input to results readout. With the aid of a smartphone vision tool, high accuracy (AUC = 1.0; CI: 0.715 - 1.00) is achieved when the CASCADE system is tested with nasopharyngeal swab samples of PCR-positive COVID-19 patients.
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