Essential properties and pitfalls of colorimetric Reverse Transcription Loop-mediated Isothermal Amplification as a point-of-care test for SARS-CoV-2 diagnosis.

Autor: de Oliveira Coelho B; Laboratory for Applied Science and Technology in Health, Carlos Chagas Institute, Oswaldo Cruz Foundation (Fiocruz), Prof Algacyr Munhoz Mader 3775 Street, Curitiba, Paraná, 81350-010, Brazil., Sanchuki HBS; Laboratory for Applied Science and Technology in Health, Carlos Chagas Institute, Oswaldo Cruz Foundation (Fiocruz), Prof Algacyr Munhoz Mader 3775 Street, Curitiba, Paraná, 81350-010, Brazil., Zanette DL; Laboratory for Applied Science and Technology in Health, Carlos Chagas Institute, Oswaldo Cruz Foundation (Fiocruz), Prof Algacyr Munhoz Mader 3775 Street, Curitiba, Paraná, 81350-010, Brazil., Nardin JM; Laboratory for Applied Science and Technology in Health, Carlos Chagas Institute, Oswaldo Cruz Foundation (Fiocruz), Prof Algacyr Munhoz Mader 3775 Street, Curitiba, Paraná, 81350-010, Brazil., Morales HMP; Erasto Gaertner Hospital, Dr. Ovande do Amaral 201 Street, Curitiba, Paraná, 81520-060, Brazil., Fornazari B; Erasto Gaertner Hospital, Dr. Ovande do Amaral 201 Street, Curitiba, Paraná, 81520-060, Brazil., Aoki MN; Laboratory for Applied Science and Technology in Health, Carlos Chagas Institute, Oswaldo Cruz Foundation (Fiocruz), Prof Algacyr Munhoz Mader 3775 Street, Curitiba, Paraná, 81350-010, Brazil. mateus.aoki@fiocruz.br., Blanes L; Laboratory for Applied Science and Technology in Health, Carlos Chagas Institute, Oswaldo Cruz Foundation (Fiocruz), Prof Algacyr Munhoz Mader 3775 Street, Curitiba, Paraná, 81350-010, Brazil. lucas.blanes@fiocruz.br.
Jazyk: angličtina
Zdroj: Molecular medicine (Cambridge, Mass.) [Mol Med] 2021 Mar 26; Vol. 27 (1), pp. 30. Date of Electronic Publication: 2021 Mar 26.
DOI: 10.1186/s10020-021-00289-0
Abstrakt: Background: SARS-CoV-2 Reverse Transcription Loop-mediated Isothermal Amplification (RT-LAMP) colorimetric detection is a sensitive and specific point-of-care molecular biology technique used to detect the virus in only 30 min. In this manuscript we have described a few nuances of the technique still not properly described in the literature: the presence of three colors clusters; the correlation of the viral load with the color change; and the importance of using an internal control to avoid false-negative results.
Methods: To achieve these findings, we performed colorimetric RT-LAMP assays of 466 SARS-CoV-2 RT-qPCR validated clinical samples, with color quantification measured at 434 nm and 560 nm.
Results: First we determinate a sensitivity of 93.8% and specificity of 90.4%. In addition to the pink (negative) and yellow (positive) produced colors, we report for the first time the presence of an orange color cluster that may lead to wrong diagnosis. We also demonstrated using RT-qPCR and RT-LAMP that low viral loads are related to Ct values > 30, resulting in orange colors. We also demonstrated that the diagnosis of COVID-19 by colorimetric RT-LAMP is efficient until the fifth symptoms day when the viral load is still relatively high.
Conclusion: This study reports properties and indications for colorimetric RT-LAMP as point-of-care for SARS-CoV-2 diagnostic, reducing false results, interpretations and optimizing molecular diagnostics tests application.
Databáze: MEDLINE
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