期刊
FRONTIERS IN CELLULAR AND INFECTION MICROBIOLOGY
卷 12, 期 -, 页码 -出版社
FRONTIERS MEDIA SA
DOI: 10.3389/fcimb.2022.856553
关键词
carryover contamination; loop-mediated isothermal amplification (LAMP); SARS-CoV-2; COVID-19; Cod uracil DNA glycosylase (Cod-UNG); on-site testing; point-of-care
资金
- Department of Biotechnology and Biomedicine, Technical University of Denmark, Denmark
- EU H2020 [101003562, 773422]
- Danish government
- COVIDTESTS
- H2020 Societal Challenges Programme [773422] Funding Source: H2020 Societal Challenges Programme
This study developed a new LAMP assay that can eliminate carryover contamination and rapidly detect SARS-CoV-2. The assay can detect the virus as low as 2 copies/μl within 45 minutes and eliminate a significant amount of contaminants. Analysis of clinical samples showed high agreement with rRT-PCR.
Loop-mediated isothermal amplification (LAMP) is being used as a robust rapid diagnostic tool to prevent the transmission of infectious diseases. However, carryover contamination of LAMP-amplified products originating from previous tests has been a problem in LAMP-based bio-analytical assays. In this study, we developed a Cod-uracil-DNA-glycosylase real-time reverse transcriptase LAMP assay (Cod-UNG-rRT-LAMP) for the elimination of carryover contamination and the rapid detection of SARS-CoV-2 in point-of-care (POC) testing. Using the Cod-UNG-rRT-LAMP assay, the SARS-CoV-2 virus could be detected as low as 2 copies/mu l (8 copies/reaction) within 45 min of amplification and 2.63 +/- 0.17 pg (equivalent to 2.296 x 10(9) copies) of contaminants per reaction could be eliminated. Analysis of clinical SARS-CoV-2 samples using the Cod-UNG-rRT-LAMP assay showed an excellent agreement with a relative accuracy of 98.2%, sensitivity of 97.1%, and specificity of 95.2% in comparison to rRT-PCR. The results obtained in this study clearly demonstrate the feasibility of the use of the Cod-UNG-rRT-LAMP assay for applications toward the POC diagnosis of SARS-CoV-2 and on-site testing of other pathogens.
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