Rapid detection of SARS-CoV-2 and other respiratory viruses by using LAMP method with Nanopore Flongle workflow

Li, Jingjing; Quan, Weipeng; Yan, Shuge; Wu, Shuangju; Qin, Jianhu; Yang, Tingting; Liang, Fan; Wang, Depeng; Yu, Liang

doi:10.1101/2020.06.03.131474

Cited by 5 publications

(4 citation statements)

References 17 publications

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“…However, the optimal temperature for most Cas enzymes deployed in diagnostic the efficacy of different sets of LAMP primers targeting the human POP7, ACTB, or GAPDH gene. The primers labelled with "Set1", "Set2", or "Set3" are newly designed, while the primers labelled with "Pub" have been published 17,72,73 . 2 µl heat-treated saliva was used as sample input to RT-LAMP, which was performed at 65°C over 40 min in a real-time instrument.…”

Section: Discussionmentioning

confidence: 99%

An engineered CRISPR-Cas12a variant and DNA-RNA hybrid guides enable robust and rapid COVID-19 testing

et al. 2021

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Extensive testing is essential to break the transmission of SARS-CoV-2, which causes the ongoing COVID-19 pandemic. Here, we present a CRISPR-based diagnostic assay that is robust to viral genome mutations and temperature, produces results fast, can be applied directly on nasopharyngeal (NP) specimens without RNA purification, and incorporates a human internal control within the same reaction. Specifically, we show that the use of an engineered AsCas12a enzyme enables detection of wildtype and mutated SARS-CoV-2 and allows us to perform the detection step with loop-mediated isothermal amplification (LAMP) at 60-65 °C. We also find that the use of hybrid DNA-RNA guides increases the rate of reaction, enabling our test to be completed within 30 minutes. Utilizing clinical samples from 72 patients with COVID-19 infection and 57 healthy individuals, we demonstrate that our test exhibits a specificity and positive predictive value of 100% with a sensitivity of 50 and 1000 copies per reaction (or 2 and 40 copies per microliter) for purified RNA samples and unpurified NP specimens respectively.

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Section: Discussionmentioning

confidence: 99%

An engineered CRISPR-Cas12a variant and DNA-RNA hybrid guides enable robust and rapid COVID-19 testing

et al. 2021

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“…Nanopore Flongle was developed to be a quick, accessible and cost‐efficient real‐time sequencing. The nanopore Flongle workflow was used for genome analysis and identification of SARS‐CoV‐2 after 30‐min sequencing 56 …”

Section: The Sars‐cov‐2 Lamp Assays Studiesmentioning

confidence: 99%

Loop‐mediated isothermal amplification (LAMP): An effective molecular point‐of‐care technique for the rapid diagnosis of coronavirus SARS‐CoV‐2

Chaouch

2021

Reviews in Medical Virology

116

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Summary The novel coronavirus disease‐2019 (Covid‐19) public health emergency has caused enormous loss around the world. This pandemic is a concrete example of the existing gap between availability of advanced diagnostics and current need for cost‐effective methodology. The advent of the loop‐mediated isothermal amplification (LAMP) assay provided an innovative tool for establishing a rapid diagnostic technique based on the molecular amplification of pathogen RNA or DNA. In this review, we explore the applications, diagnostic effectiveness of LAMP test for molecular diagnosis and surveillance of severe acute respiratory syndrome coronavirus 2. Our results show that LAMP can be considered as an effective point‐of‐care test for the diagnosis of Covid‐19 in endemic areas, especially for low‐ and middle‐income countries.

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Section: Viral Nucleic Acid Testsmentioning

confidence: 99%

“…88 Li et al demonstrated a combined LAMP-Nanopore Flongle real-time sequencing workflow, wherein COVID-19 RNA was amplified using LAMP for 30 min, prior to being transferred to the sequencing element. The combined approach has an LOD of 21.2 × 10 3 copies/mL (212 copies/reaction), which can be performed in under 2 h. 86 These examples demonstrate the potential of utilizing portable sequencing methods in POC diagnostics; however, these need to be combined with nucleic acid amplification for reaching the sensitivity needed for clinical diagnostics and are often faced with challenges related to clogging when interfaced with raw biological samples. 89…”

Section: ■ Viral Nucleic Acid Testsmentioning

confidence: 99%

Roadmap to the Bioanalytical Testing of COVID-19: From Sample Collection to Disease Surveillance

et al. 2020

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The disease caused by SARS-CoV-2, coronavirus disease 2019 (COVID-19), has led to a global pandemic with tremendous mortality, morbidity, and economic loss. The current lack of effective vaccines and treatments places tremendous value on widespread screening, early detection, and contact tracing of COVID-19 for controlling its spread and minimizing the resultant health and societal impact. Bioanalytical diagnostic technologies have played a critical role in the mitigation of the COVID-19 pandemic and will continue to be foundational in the prevention of the subsequent waves of this pandemic along with future infectious disease outbreaks. In this Review, we aim at presenting a roadmap to the bioanalytical testing of COVID-19, with a focus on the performance metrics as well as the limitations of various techniques. The state-of-the-art technologies, mostly limited to centralized laboratories, set the clinical metrics against which the emerging technologies are measured. Technologies for point-of-care and do-it-yourself testing are rapidly emerging, which open the route for testing in the community, at home, and at points-of-entry to widely screen and monitor individuals for enabling normal life despite of an infectious disease pandemic. The combination of different classes of diagnostic technologies (centralized and point-of-care and relying on multiple biomarkers) are needed for effective diagnosis, treatment selection, prognosis, patient monitoring, and epidemiological surveillance in the event of major pandemics such as COVID-19.

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Rapid detection of SARS-CoV-2 and other respiratory viruses by using LAMP method with Nanopore Flongle workflow

Cited by 5 publications

References 17 publications

An engineered CRISPR-Cas12a variant and DNA-RNA hybrid guides enable robust and rapid COVID-19 testing

An engineered CRISPR-Cas12a variant and DNA-RNA hybrid guides enable robust and rapid COVID-19 testing

Loop‐mediated isothermal amplification (LAMP): An effective molecular point‐of‐care technique for the rapid diagnosis of coronavirus SARS‐CoV‐2

Roadmap to the Bioanalytical Testing of COVID-19: From Sample Collection to Disease Surveillance

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