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Article

Highly Specific and Rapid Detection of Hepatitis C Virus Using RT-LAMP-Coupled CRISPR–Cas12 Assay

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Division of Clinical Microbiology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand
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Maladies Infectieuses et Vecteurs: Écologie, Génétique, Évolution et Contrôle (MIVEGEC), Agropolis University Montpellier, Centre National de la Recherche Scientifique (CNRS), Institut de Recherche Pour le Développement (IRD), 34394 Montpellier, France
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Associated Medical Sciences (AMS)-PHPT Research Collaboration, Chiang Mai 50200, Thailand
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Infectious Diseases Research Unit, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand
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Research Institute for Health Sciences, Chiang Mai University, Chiang Mai 50200, Thailand
*
Authors to whom correspondence should be addressed.
Academic Editor: Nuno Taveira
Diagnostics 2022, 12(7), 1524; https://doi.org/10.3390/diagnostics12071524
Received: 27 May 2022 / Revised: 17 June 2022 / Accepted: 17 June 2022 / Published: 23 June 2022
(This article belongs to the Collection Diagnostic Virology)
Hepatitis C virus (HCV) infection can be cured with pan-genotypic direct-acting antiviral agents. However, identifying individuals with current hepatitis C remains a major challenge, especially in resource-limited settings where access to or availability of molecular tests is still limited. The goal of this study was to develop and validate a molecular assay for the rapid detection of HCV RNA in resource-limited settings. It is based on a combination of reverse transcription loop-mediated isothermal amplification (RT-LAMP) with the clustered regularly interspaced short palindromic repeats–CRISPR-associated protein 12a (CRISPR–Cas12a) cleavage assay that allows the recognition of specific HCV nucleic acid sequences. Amplified products after the cleavage reactions can be visualized on lateral flow strips or measured with a fluorescence detector. When tested on clinical samples from individuals infected with HCV, HIV, or HBV, or from healthy donors, the RT-LAMP-coupled CRISPR–Cas12 assay yielded 96% sensitivity, 100% specificity, and 97% agreement as compared to the reference method (Roche COBAS AmpliPrep/COBAS TaqMan HCV Test). This assay could detect HCV RNA concentrations as low as 10 ng/µL (an estimated 2.38 Log10 IU/mL). Therefore, this sensitive and specific assay may represent an affordable and reliable point-of-care test for the identification of individuals with active hepatitis C in low-resource settings. View Full-Text
Keywords: hepatitis C virus; HCV RNA; chronic hepatitis C; point-of-care testing; RT-LAMP; CRISPR–Cas12; lateral flow-based assay; fluorescence-based assay hepatitis C virus; HCV RNA; chronic hepatitis C; point-of-care testing; RT-LAMP; CRISPR–Cas12; lateral flow-based assay; fluorescence-based assay
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MDPI and ACS Style

Kham-Kjing, N.; Ngo-Giang-Huong, N.; Tragoolpua, K.; Khamduang, W.; Hongjaisee, S. Highly Specific and Rapid Detection of Hepatitis C Virus Using RT-LAMP-Coupled CRISPR–Cas12 Assay. Diagnostics 2022, 12, 1524. https://doi.org/10.3390/diagnostics12071524

AMA Style

Kham-Kjing N, Ngo-Giang-Huong N, Tragoolpua K, Khamduang W, Hongjaisee S. Highly Specific and Rapid Detection of Hepatitis C Virus Using RT-LAMP-Coupled CRISPR–Cas12 Assay. Diagnostics. 2022; 12(7):1524. https://doi.org/10.3390/diagnostics12071524

Chicago/Turabian Style

Kham-Kjing, Nang, Nicole Ngo-Giang-Huong, Khajornsak Tragoolpua, Woottichai Khamduang, and Sayamon Hongjaisee. 2022. "Highly Specific and Rapid Detection of Hepatitis C Virus Using RT-LAMP-Coupled CRISPR–Cas12 Assay" Diagnostics 12, no. 7: 1524. https://doi.org/10.3390/diagnostics12071524

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