Tweet
Sci Rep
. 2021 Aug 10;11(1):16193.
doi: 10.1038/s41598-021-95607-1.
Clinical validation of optimised RT-LAMP for the diagnosis of SARS-CoV-2 infection
Boon Lim # 1 2 , Jeremy Ratcliff # 3 , Dorota A Nawrot # 4 , Yejiong Yu 1 2 , Harshmeena R Sanghani 4 , Chia-Chen Hsu 1 2 , Leon Peto 5 , Simon Evans 5 , Susanne H Hodgson 5 6 , Aikaterini Skeva 7 , Maria Adam 7 , Maria Panopoulou 7 , Christos E Zois 7 , Katy Poncin 8 , Sridhar R Vasudevan 4 , Siqi Dai 1 2 , Shuai Ren 1 2 , Hong Chang 9 , Zhanfeng Cui 1 2 9 , Peter Simmonds 3 , Wei E Huang 10 11 , Monique I Andersson 12 13
Affiliations
- PMID: 34376716
- DOI: 10.1038/s41598-021-95607-1
Abstract
We have optimised a reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay for the detection of SARS-CoV-2 from extracted RNA for clinical application. We improved the stability and reliability of the RT-LAMP assay by the addition of a temperature-dependent switch oligonucleotide to reduce self- or off-target amplification. We then developed freeze-dried master mix for single step RT-LAMP reaction, simplifying the operation for end users and improving long-term storage and transportation. The assay can detect as low as 13 copies of SARS-CoV2 RNA per reaction (25-μL). Cross reactivity with other human coronaviruses was not observed. We have applied the new RT-LAMP assay for testing clinical extracted RNA samples extracted from swabs of 72 patients in the UK and 126 samples from Greece and demonstrated the overall sensitivity of 90.2% (95% CI 83.8-94.7%) and specificity of 92.4% (95% CI 83.2-97.5%). Among 115 positive samples which Ct values were less than 34, the RT-LAMP assay was able to detect 110 of them with 95.6% sensitivity. The specificity was 100% when RNA elution used RNase-free water. The outcome of RT-LAMP can be reported by both colorimetric detection and quantifiable fluorescent reading. Objective measures with a digitized reading data flow would allow for the sharing of results for local or national surveillance.