tetano
Editor, Senior Moderator
Anal Chim Acta
. 2023 May 15;1255:341146.
doi: 10.1016/j.aca.2023.341146. Epub 2023 Mar 25.
A triple-target reverse transcription loop-mediated isothermal amplification (RT-LAMP) for rapid and accurate detection of SARS-CoV-2 virus
Cong Zhang[SUP] 1 [/SUP], Ji Lv[SUP] 1 [/SUP], Yanan Cao[SUP] 1 [/SUP], Xiaowei Yao[SUP] 1 [/SUP], Mingzhu Yin[SUP] 1 [/SUP], Shuiqing Li[SUP] 1 [/SUP], Junping Zheng[SUP] 2 [/SUP], Hongtao Liu[SUP] 3 [/SUP]
Affiliations
Abstract
The spreading of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) across the world has impacted people's health and lives worldwide in recent years. Rapid and accurate diagnosis is crucial for curbing the pandemic of coronavirus disease 2019 (COVID-19). Reverse transcription loop-mediated isothermal amplification (RT-LAMP) has great potential for SARS-CoV-2 detection but fails to completely replace conventional PCR due to the high false-positive rate (FPR). We proposed a triple-target RT-LAMP method for dual-signal, sensitive, and simultaneous detection of conserved genes of SARS-CoV-2. Multiple LAMP primer sets were designed for N, E, and M genes and their amplification efficacy were screened. Then, using artificial plasmids and RNA, the optimal primer set for each gene was examined on specificity, sensitivity, and detection range. The RT-LAMP initiated by these primer sets exhibited better specificity and sensitivity than that of RT-qPCR, and the triple-target RT-LAMP could determine different variants of SARS-CoV-2. By testing 78 artificial RNA samples, the total FPR of triple-target RT-LAMP was eliminated compared with that of mono-target RT-LAMP. The triple-target RT-LAMP method precisely identified throat swab specimens through colorimetry and fluorescent signals within 60 min, and the limit of detection (LOD) was as low as 187 copies/reaction. In the future, the triple-target RT-LAMP can be applied to in-field and on-site diagnosis of symptomatic and asymptomatic virus carriers.
Keywords: COVID-19; Colorimetry; LAMP; Multi-target; SARS-CoV-2.
. 2023 May 15;1255:341146.
doi: 10.1016/j.aca.2023.341146. Epub 2023 Mar 25.
A triple-target reverse transcription loop-mediated isothermal amplification (RT-LAMP) for rapid and accurate detection of SARS-CoV-2 virus
Cong Zhang[SUP] 1 [/SUP], Ji Lv[SUP] 1 [/SUP], Yanan Cao[SUP] 1 [/SUP], Xiaowei Yao[SUP] 1 [/SUP], Mingzhu Yin[SUP] 1 [/SUP], Shuiqing Li[SUP] 1 [/SUP], Junping Zheng[SUP] 2 [/SUP], Hongtao Liu[SUP] 3 [/SUP]
Affiliations
- PMID: 37032059
- DOI: 10.1016/j.aca.2023.341146
Abstract
The spreading of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) across the world has impacted people's health and lives worldwide in recent years. Rapid and accurate diagnosis is crucial for curbing the pandemic of coronavirus disease 2019 (COVID-19). Reverse transcription loop-mediated isothermal amplification (RT-LAMP) has great potential for SARS-CoV-2 detection but fails to completely replace conventional PCR due to the high false-positive rate (FPR). We proposed a triple-target RT-LAMP method for dual-signal, sensitive, and simultaneous detection of conserved genes of SARS-CoV-2. Multiple LAMP primer sets were designed for N, E, and M genes and their amplification efficacy were screened. Then, using artificial plasmids and RNA, the optimal primer set for each gene was examined on specificity, sensitivity, and detection range. The RT-LAMP initiated by these primer sets exhibited better specificity and sensitivity than that of RT-qPCR, and the triple-target RT-LAMP could determine different variants of SARS-CoV-2. By testing 78 artificial RNA samples, the total FPR of triple-target RT-LAMP was eliminated compared with that of mono-target RT-LAMP. The triple-target RT-LAMP method precisely identified throat swab specimens through colorimetry and fluorescent signals within 60 min, and the limit of detection (LOD) was as low as 187 copies/reaction. In the future, the triple-target RT-LAMP can be applied to in-field and on-site diagnosis of symptomatic and asymptomatic virus carriers.
Keywords: COVID-19; Colorimetry; LAMP; Multi-target; SARS-CoV-2.