CRISPR-ENHANCE: An enhanced nucleic acid detection platform using Cas12a

Methods. 2022 Jul;203:116-124. doi: 10.1016/j.ymeth.2021.02.001. Epub 2021 Feb 9.


Rapid detection of nucleic acids is essential for clinical diagnosis of a wide range of infectious and non-infectious diseases. CRISPR-based diagnostic platforms are well-established for rapid and specific detection of nucleic acids but suffer from a low detection sensitivity without a target pre-amplification step. Our recently developed detection system, called CRISPR-ENHANCE, employs engineered crRNAs and optimized conditions to achieve a significantly higher sensitivity and enable femtomolar levels of nucleic acid detection even without target pre-amplification. Using the CRISPR-ENHANCE platform and following the methodology detailed in this paper, nucleic acid detection for low copy numbers can be achieved in less than an hour through either a fluorescence-based detection or a lateral flow assay. The step-by-step instructions provided, in addition to describing how to perform both assays, incorporate details on a LAMP/RT-LAMP-based target amplification step to enable detection of RNA, ssDNA and dsDNA. Furthermore, a protocol for in-house expression and purification of LbCas12a using CL7/lm7-based affinity chromatography, which has been used to achieve a high yield and purity of the enzyme in a single-step, is provided.

Keywords: CL7; COVID-19; CRISPR; CRISPR/Cas; Cas12a; DETECTR; Detection; Diagnostics; ENHANCE; Fluorescence; Lateral Flow Assay; Nucleic Acids; Protein purification; RT-LAMP; SARS-CoV-2.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CRISPR-Cas Systems / genetics
  • DNA, Single-Stranded / genetics
  • Nucleic Acid Amplification Techniques / methods
  • Nucleic Acids*
  • SARS-CoV-2*


  • DNA, Single-Stranded
  • Nucleic Acids