Standard operating procedures for SARS-CoV-2 detection by a clinical diagnostic RT-LAMP assay

Buck, Michael D.; Poirier, Enzo Z.; Cardoso, Ana; Frederico, Bruno; Canton, Johnathan; Barrell, Sam; Beale, Rupert; Byrne, Richard; Caidan, Simon; Crawford, Margaret; Cubitt, Laura; Gamblin, Steve; Gandhi, Sonia; Goldstone, Robert; Grant, Paul; Gulati, Kiran; Hindmarsh, Steve; Howell, Michael; Hubank, Michael; Instrell, Rachael; Jiang, Ming; Kassiotis, George; Lu, Wei-Ting; MacRae, James I.; Martini, Iana; Miller, Davin; Moore, D. J.; Nastouli, Eleni; Nicod, Jérôme; Nightingale, Luke; Olsen, Jessica; Oomatia, Amin; O’Reilly, Nicola; Rideg, Anett; Song, Ok‐Ryul; Strange, Amy; Swanton, Charles; Turajlic, Samra; Walker, P.A.; Wu, Mary; Sousa, Caetano Reis e

doi:10.1101/2020.06.29.20142430

Cited by 13 publications

(16 citation statements)

References 15 publications

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“…As a result, some samples might give a positive result to the antibody test but www.nature.com/scientificreports/ provide a negative result to the LAMP test. Buck et al (2020), Thi et al (2020) and Rodel et al (2020) are also studies with sensitivity below 80% 34,39,72 . These studies also reported quantity of viral RNA (as Ct value of RT-qPCR) in purified RNA sample.…”

Section: Sensitivity Specificity and Diagnostic Odd Ratio (Dor) Of Nmentioning

confidence: 99%

The diagnostic accuracy of isothermal nucleic acid point-of-care tests for human coronaviruses: A systematic review and meta-analysis

Subsoontorn

Lohitnavy

Kongkaew

2020

Sci Rep

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Many recent studies reported coronavirus point-of-care tests (POCTs) based on isothermal amplification. However, the performances of these tests have not been systematically evaluated. Cochrane Handbook for Systematic Reviews of Diagnostic Test Accuracy was used as a guideline for conducting this systematic review. We searched peer-reviewed and preprint articles in PubMed, BioRxiv and MedRxiv up to 28 September 2020 to identify studies that provide data to calculate sensitivity, specificity and diagnostic odds ratio (DOR). Quality Assessment of Diagnostic Accuracy Studies 2 (QUADAS-2) was applied for assessing quality of included studies and Preferred Reporting Items for a Systematic Review and Meta-analysis of Diagnostic Test Accuracy Studies (PRISMA-DTA) was followed for reporting. We included 81 studies from 65 research articles on POCTs of SARS, MERS and COVID-19. Most studies had high risk of patient selection and index test bias but low risk in other domains. Diagnostic specificities were high (> 0.95) for included studies while sensitivities varied depending on type of assays and sample used. Most studies (n = 51) used reverse transcription loop-mediated isothermal amplification (RT-LAMP) to diagnose coronaviruses. RT-LAMP of RNA purified from COVID-19 patient samples had pooled sensitivity at 0.94 (95% CI: 0.90–0.96). RT-LAMP of crude samples had substantially lower sensitivity at 0.78 (95% CI: 0.65–0.87). Abbott ID Now performance was similar to RT-LAMP of crude samples. Diagnostic performances by CRISPR and RT-LAMP on purified RNA were similar. Other diagnostic platforms including RT- recombinase assisted amplification (RT-RAA) and SAMBA-II also offered high sensitivity (> 0.95). Future studies should focus on the use of un-bias patient cohorts, double-blinded index test and detection assays that do not require RNA extraction.

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Section: Sensitivity Specificity and Diagnostic Odd Ratio (Dor) Of Nmentioning

confidence: 99%

The diagnostic accuracy of isothermal nucleic acid point-of-care tests for human coronaviruses: A systematic review and meta-analysis

Subsoontorn

Lohitnavy

Kongkaew

2020

Sci Rep

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“…February and 8th April 2020, six independent groups have posted preprints of submitted manuscripts evaluating novel RT-LAMP testing methods against RT-PCR as gold standard (Table 1). Since then, a number [13] of other groups have published high-quality studies demonstrating that RT-LAMP has the potential to replace RT-PCR as a means for detecting SARS-CoV-2 (Severe acute respiratory syndrome coronavirus 2) within RNA extracted from nose -throat swabs and endotracheal secretions/bronchoalveolar lavage fluid [5,14,15].…”

Section: Introductionmentioning

confidence: 99%

Detecting SARS-CoV-2 at point of care: preliminary data comparing loop-mediated isothermal amplification (LAMP) to polymerase chain reaction (PCR)

et al. 2020

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Background A cost effective and efficient diagnostic tool for COVID-19 as near to the point of care (PoC) as possible would be a game changer in the current pandemic. We tested reverse transcription loop mediated isothermal amplification (RT-LAMP), a method which can produce results in under 30 min, alongside standard methods in a real-life clinical setting. Methods This prospective service improvement project piloted an RT-LAMP method on nasal and pharyngeal swabs on 21 residents of a high dependency care home, with two index COVID-19 cases, and compared it to multiplex tandem reverse transcription polymerase chain reaction (RT-PCR). We recorded vital signs of patients to correlate clinical and laboratory information and calculated the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of a single swab using RT-LAMP compared with the current standard, RT-PCR, as per Standards for Reporting Diagnostic Accuracy Studies (STARD) guidelines. Results The novel method accurately detected 8/10 RT-PCR positive cases and identified a further 3 positive cases. Eight further cases were negative using both methods. Using repeated RT-PCR as a “gold standard”, the sensitivity and specificity of a single novel test were 80 and 73% respectively. PPV was 73% and NPV was 83%. Incorporating retesting of low signal RT-LAMP positives improved the specificity to 100%. We also speculate that hypothermia may be a significant early clinical sign of COVID-19. Conclusions RT-LAMP testing for SARS-CoV-2 was found to be promising, fast and to work equivalently to RT-PCR methods. RT-LAMP has the potential to transform COVID-19 detection, bringing rapid and accurate testing to the PoC. RT-LAMP could be deployed in mobile community testing units, care homes and hospitals to detect disease early and prevent spread.

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“…A scheme for evaluating the results is shown in Supp. Table 2. Due to the removal of non-target nucleic acids and the inclusion of LAMP enhancing enzymes in Cap-iLAMP, we did not detect false positive results, which were often reported in previous LAMP-based detection methods (Teng et al 2007;Abbasi et al 2016 preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in The copyright holder for this this version posted August 6, 2020. ; https://doi.org/10.1101/2020.08.04.20168617 doi: medRxiv preprint CoV-2 detection methods (Broughton et al 2020;Buck et al 2020;Corman et al 2020;Dao Thi et al 2020) in some key aspects as shown in Table 1.…”

Section: Discussionmentioning

confidence: 99%

Rapid, reliable, and cheap point-of-care bulk testing for SARS-CoV-2 by combining hybridization capture with improved colorimetric LAMP (Cap-iLAMP)

Bokelmann

Nickel

Maričić

et al. 2020

Preprint

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Efforts to contain the spread of SARS-CoV-2 have spurred the need for reliable, rapid, and cost-effective diagnostic methods which can easily be applied to large numbers of people. However, current standard protocols for the detection of viral nucleic acids while sensitive, require a high level of automation, sophisticated laboratory equipment and trained personnel to achieve throughputs that allow whole communities to be tested on a regular basis. Here we present Cap-iLAMP (capture and improved loop-mediated isothermal amplification). This method combines a hybridization capture-based RNA extraction of non-invasive gargle lavage samples to concentrate samples and remove inhibitors with an improved colorimetric RT-LAMP assay and smartphone-based color scoring. Cap-iLAMP is compatible with point-of-care testing and enables the detection of SARS-CoV-2 positive samples in less than one hour. In contrast to direct addition of the sample to improved LAMP (iLAMP), Cap-iLAMP does not result in false positives and single infected samples can be detected in a pool among 25 uninfected samples, thus reducing the technical cost per test to ~1 Euro per individual.

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Standard operating procedures for SARS-CoV-2 detection by a clinical diagnostic RT-LAMP assay

Cited by 13 publications

References 15 publications

The diagnostic accuracy of isothermal nucleic acid point-of-care tests for human coronaviruses: A systematic review and meta-analysis

The diagnostic accuracy of isothermal nucleic acid point-of-care tests for human coronaviruses: A systematic review and meta-analysis

Detecting SARS-CoV-2 at point of care: preliminary data comparing loop-mediated isothermal amplification (LAMP) to polymerase chain reaction (PCR)

Rapid, reliable, and cheap point-of-care bulk testing for SARS-CoV-2 by combining hybridization capture with improved colorimetric LAMP (Cap-iLAMP)

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