CRISPR-Based Electrochemical Sensor Permits Sensitive and SpecificViral Detection in Low-Resource Settings

摘要

As shown in Figure ​Figure11, the electrochemical sensor is used to detectviral genetic material in a three-step process. Traditional methodsare first used to extract viral genetic material, which is then amplifiedusing loop-mediated isothermal amplification (LAMP)—an isothermalmethod that is feasible to implement in low-resource settings. LAMPproducts are added to a CRISPR-based recognition system that is activatedin the presence of target amplicon, circumventing the challenge ofnonspecific amplification associated with LAMP.3 The Cas12a reaction mix is deposited on gold electrodesthat are functionalized with methylene blue (MB) tagged oligonucleotides(Figure ​Figure22). The gRNAof Cas12a is engineered so that specific loci in each viral genomeof interest activate the endonuclease and cause it to cleave the MB-taggedoligonucleotide, thereby resulting in an electrochemical signal. Electrochemicalsignal from MB is measured before and after treatment with the Cas12aenzyme to detect whether the virus is present.