- SHERLOCK (Specific High-sensitivity Enzymatic Reporter unLOCKing) is a nucleic acid detection platform that leverages the collateral cleavage activity of CRISPR-associated enzymes, particularly Cas13a, for highly sensitive, specific, and rapid detection of DNA and RNA. Developed by the Feng Zhang lab at the Broad Institute, SHERLOCK represents a significant advancement in molecular diagnostics, enabling the detection of low-abundance nucleic acid sequences without the need for traditional amplification-based assays.
- At the core of SHERLOCK is the RNA-guided enzyme Cas13a (formerly known as C2c2), which becomes activated upon binding to its target RNA sequence via a CRISPR RNA (crRNA). Once activated, Cas13a exhibits collateral cleavage activity, meaning it nonspecifically cleaves nearby single-stranded RNA molecules in addition to the specific target. This unique property is harnessed in SHERLOCK by introducing a reporter RNA molecule labeled with a fluorophore and a quencher. Upon activation of Cas13a by the target RNA, the reporter is cleaved, leading to the release of fluorescence, which serves as a detectable signal.
- To achieve extremely high sensitivity, SHERLOCK incorporates isothermal amplification steps such as recombinase polymerase amplification (RPA) or loop-mediated isothermal amplification (LAMP) prior to CRISPR detection. These amplification steps increase the number of target molecules at a constant temperature, making the platform suitable for point-of-care and resource-limited settings. In addition, SHERLOCK can be easily adapted to detect DNA targets by first converting them into RNA using T7 transcription.
- SHERLOCK’s specificity arises from the programmable nature of the CRISPR-Cas system, which allows it to discriminate between sequences that differ by as little as a single nucleotide. This makes the platform particularly valuable for applications such as pathogen detection, SNP genotyping, and detection of drug resistance mutations. Its adaptability and high sensitivity (down to attomolar concentrations) have made it a leading technology in infectious disease diagnostics, including for Zika virus, Dengue virus, and SARS-CoV-2.
- Moreover, the SHERLOCK platform has been expanded into multiplexed formats (e.g., SHERLOCKv2), lateral flow readouts, and paper-based diagnostics, increasing its utility in field settings. These developments have made SHERLOCK a cornerstone in the CRISPR-based diagnostic toolkit, offering a powerful, flexible, and low-cost alternative to conventional PCR and antibody-based methods.
