An amplification-free ultra-sensitive electrochemical CRISPR/Cas biosensor for drug-resistant bacteria detection.

Journal article

Suea-Ngam, A., Howes, P. and deMello, A. (2021). An amplification-free ultra-sensitive electrochemical CRISPR/Cas biosensor for drug-resistant bacteria detection. Chemical science. 12 (38), pp. 12733-12743. https://doi.org/10.1039/d1sc02197d
AuthorsSuea-Ngam, A., Howes, P. and deMello, A.
AbstractContinued development of high-performance and cost-effective diagnostic tools is vital for improving infectious disease treatment and transmission control. For nucleic acid diagnostics, moving beyond enzyme-mediated amplification assays will be critical in reducing the time and complexity of diagnostic technologies. Further, an emerging area of threat, in which diagnostics will play an increasingly important role, is antimicrobial resistance (AMR) in bacterial infections. Herein, we present an amplification-free electrochemical CRISPR/Cas biosensor utilizing silver metallization (termed E-Si-CRISPR) to detect methicillin-resistant (MRSA). Using a custom-designed guide RNA (gRNA) targeting the gene of MRSA, the Cas12a enzyme allows highly sensitive and specific detection when employed with silver metallization and square wave voltammetry (SWV). Our biosensor exhibits excellent analytical performance, with detection and quantitation limits of 3.5 and 10 fM, respectively, and linearity over five orders of magnitude (from 10 fM to 0.1 nM). Importantly, we observe no degradation in performance when moving from buffer to human serum samples, and achieve excellent selectivity for MRSA in human serum in the presence of other common bacteria. The E-Si-CRISPR method shows significant promise as an ultrasensitive field-deployable device for nucleic acid-based diagnostics, without requiring nucleic acid amplification. Finally, adjustment to a different disease target can be achieved by simple modification of the gRNA protospacer. [Abstract copyright: This journal is © The Royal Society of Chemistry.]
Year2021
JournalChemical science
Journal citation12 (38), pp. 12733-12743
PublisherRoyal Society of Chemistry (RSC)
ISSN2041-6520
Digital Object Identifier (DOI)https://doi.org/10.1039/d1sc02197d
Publication dates
Online03 Sep 2021
Publication process dates
Accepted13 Aug 2021
Deposited26 Jan 2022
Accepted author manuscript
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Open
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