Amplification-free detection of SARS-CoV-2 with CRISPR-Cas13a and mobile phone microscopy

Abstract

The December 2019 outbreak of a novel respiratory virus, SARS-CoV-2, has become an ongoing global pandemic due in part to the challenge of identifying symptomatic, asymptomatic, and pre-symptomatic carriers of the virus. CRISPR diagnostics can augment gold-standard PCR-based testing if they can be made rapid, portable, and accurate. Here, we report the development of an amplification-free CRISPR-Cas13a assay for direct detection of SARS-CoV-2 from nasal swab RNA that can be read with a mobile phone microscope. The assay achieved ∼100 copies/μL sensitivity in under 30 min of measurement time and accurately detected pre-extracted RNA from a set of positive clinical samples in under 5 min. We combined crRNAs targeting SARS-CoV-2 RNA to improve sensitivity and specificity and directly quantified viral load using enzyme kinetics. Integrated with a reader device based on a mobile phone, this assay has the potential to enable rapid, low-cost, point-of-care screening for SARS-CoV-2.

Graphical Abstract

Highlights

• •

  CRISPR-Cas13a can quantitatively detect SARS-CoV-2 RNA without pre-amplification

• •

  Combining crRNAs targeting multiple regions of the viral RNA enhances sensitivity

• •

  Cas13a can accurately and rapidly quantify SARS-CoV-2 RNA in patient samples

• •

  A mobile phone-based device allows for portable and sensitive readout of the assay

Abstract

Fozouni et al. devise a way to use CRISPR-Cas13a to detect and quantify SARS-CoV-2 RNA from patient samples without the need for a pre-amplification step. They then show how the assay’s signal can be efficiently detected with a portable, mobile phone-based device.