CRISPR-Based Approaches for Efficient and Accurate Detection of SARS-CoV-2

Wancun Zhang, PhD; Kangbo Liu, MD; Pin Zhang, MD; Weyland Cheng, PhD; Linfei Li, MD; Fan Zhang, MD; Zhidan Yu, PhD; Lifeng Li, PhD; Xianwei Zhang, PhD

Disclosures

Lab Med. 2021;52(2):116-121. 

In This Article

Ultra-sensitive Detection of SARS-CoV-2

Clinical studies have shown that the viral titers of hospitalized patients can fluctuate day-to-day with no correlation with the severity of the disease.[8,36,37] In 24 various specimens from patients in the recovery period, RNA was detected as negative for both the N gene and the ORF1b gene at several days after their readmission to the hospital using a commercial kit whose lower limit of detection (LOD) was relatively high (500 copies/mL). However, using a higher-sensitivity SHERLOCK kit with an LOD of 100 copies/mL, 75% of specimens were positive for the S gene and 41.6% for ORF genes, suggesting that the carrier status of the virus may exist in patients who have recovered from COVID-19.[38–42] Therefore, more sensitive RNA detection methods are required to detect and monitor these patients.

The CRISPR-nCoV approach developed by Hou et al[11] showed near single-copy sensitivity for SARS-CoV-2 detection and great clinical sensitivity with a shorter turnaround time than RT-PCR. Broughton et al[16] developed the SARS-CoV-2 DETECTR lateral flow assay, which performs simultaneous reverse transcription and isothermal amplification using LAMP followed by Cas12 detection, where it can be visualized on a lateral flow strip with a limit of detection of 10 copies/μL within 30 minutes. The CRISPR-based SHERLOCK technique for the detection of COVID-19 developed by F. Zhang et al[25] can detect COVID-19 target sequences in a range between 20 and 200 aM (10–100 copies per μL of input) within 60 minutes using synthetic COVID-19 virus RNA fragments. These ultrasensitive CRISPR-based approaches, along with STOPCovid,[27] the AIOD-CRISPR assay,[35] and the Cas12a-based detection system,[26] can accurately and effectively monitor and manage patients with COVID-19 during their recovery period.

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