Development and Validation of Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP) as a Simple and Rapid Diagnostic Tool for SARS-CoV-2 Detection

Ahmad M. Aldossary, Essam A. Tawfik, Musaad A. Altammami, Azzam A. Alquait, Rayan Y. Booq, Bandar K. Sendy, Mohammed S. Alarawi, Takashi Gojobori, Asmaa M. Altamimi, Taghreed A. Alaifan, Ahmed M. Albarrag, Essam J. Alyamani*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

Since the COVID-19 pandemic outbreak in the world, many countries have searched for quick diagnostic tools to detect the virus. There are many ways to design diagnostic assays; however, each may have its limitations. A quick, sensitive, specific, and simple approach is essential for highly rapidly transmitted infections, such as SARS-CoV-2. This study aimed to develop a rapid and cost-effective diagnostic tool using a one-step Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP) approach. The results were observed using the naked eye within 30–60 min using turbidity or colorimetric analysis. The sensitivity, specificity, and lowest limit of detection (LoD) for SARS-CoV-2 RNA against the RT-LAMP assay were assessed. This assay was also verified and validated against commercial quantitative RT-PCR used by health authorities in Saudi Arabia. Furthermore, a quick and direct sampling from the saliva, or buccal cavity, was applied after simple modification, using proteinase K and heating at 98 °C for 5 min to avoid routine RNA extraction. This rapid single-tube diagnostic tool detected COVID-19 with an accuracy rate of 95% for both genes (ORF1a and N) and an LoD for the ORF1a and N genes as 39 and 25 copies/reaction, respectively. It can be potentially used as a high-throughput national screening for different respiratory-based infections within the Middle East region, such as the MERS virus or major zoonotic pathogens such as Mycobacterium paratuberculosis and Brucella spp., particularly in remote and rural areas where lab equipment is limited.

Original languageEnglish (US)
Article number2232
JournalDiagnostics
Volume12
Issue number9
DOIs
StatePublished - Sep 2022

Keywords

  • colorimetric assay
  • COVID-19
  • N gene
  • ORF1a gene
  • Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP)
  • SARS-CoV-2

ASJC Scopus subject areas

  • Clinical Biochemistry

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