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. 2023 Jul 11;13(14):2344.
doi: 10.3390/diagnostics13142344.

Extraction-Free Colorimetric RT-LAMP Detection of SARS-CoV-2 in Saliva

Ane Rivas-Macho  1   2 , Ane Sorarrain  3 , José M Marimón  3 , Felipe Goñi-de-Cerio  1 , Garbiñe Olabarria  1
Affiliations

Extraction-Free Colorimetric RT-LAMP Detection of SARS-CoV-2 in Saliva

Ane Rivas-Macho et al. Diagnostics (Basel). .

Abstract

The pandemic situation caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has highlighted the need of fast, simple, and cost-effective tests for the diagnosis of emerging pathogens. RT-qPCR has been established as the reference technique for the diagnosis of SARS-CoV-2 infections. This method requires a time-consuming protocol for the extraction of the nucleic acids present in the sample. A colorimetric reverse transcription loop-mediated isothermal amplification using the calcein molecule combined with a simple extraction-free method for saliva samples (calcein RT-LAMP) has been developed. Samples are heated 95 °C for 10 min before amplification at 63 °C for 40 min. The results can be observed by fluorescence or by the naked eye with a color change from orange to green. The method was compared with commercialized available colorimetric and fluorescent RT-LAMP kits. The developed method shows better sensitivity and specificity than the colorimetric commercial RT-LAMP and the same as the fluorescent RT-LAMP, without the need of a fluorescent reader. Moreover, the calcein RT-LAMP has, compared to RT-qPCR, a sensitivity of 90% and a specificity of 100% for saliva samples with a Ct ≤ 34, without the need for expensive RT-qPCR instruments, demonstrating the potential of this method for population screening.

Keywords: RT-LAMP; SARS-CoV-2; calcein; extraction-free; saliva.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Colorimetric results of the specificity assay against different related coronaviruses and other respiratory viruses obtained with IH-C. Positive SARS-CoV-2 samples are in green and negative samples are in orange.
Figure 2
Figure 2
Sensitivity results of the developed WS-C with saliva samples. Positive results in green and negative ones in orange. CP: Positive control. CN: Negative control.
Figure 3
Figure 3
Sensitivity comparation of the developed extraction-free WS-C against a commercial colorimetric RT-LAMP (WS-PR) and commercial fluorescent RT-LAMP (WS-F). CN: Negative control. CP: Positive Control. nd: not detected.
See this image and copyright information in PMC

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References

    1. Mohammed F.S., Uysal I., Sevindik M. A Review on Antiviral Plants Effective against Different Virus Types. Prospect. Pharm. Sci. 2023;21:1–21. doi: 10.56782/pps.128. - DOI
    1. Coronavirus Disease (COVID-19) Pandemic. [(accessed on 5 July 2022)]. Available online: https://www.who.int/europe/emergencies/situations/covid-19.
    1. World Health Organization . Laboratory Testing for Coronavirus Disease (COVID-19) in Suspected Human Cases: Interim Guidance. World Health Organization; Geneva, Switzerland: 2020. pp. 1–7.
    1. Zhang H., Xu Y., Fohlerova Z., Chang H., Iliescu C., Neuzil P. LAMP-on-a-chip: Revising microfluidic platforms for loop-mediated DNA amplification. TrAC Trends Anal. Chem. 2019;113:44–53. doi: 10.1016/j.trac.2019.01.015. - DOI - PMC - PubMed
    1. Song Q., Sun X., Dai Z., Gao Y., Gong X., Zhou B., Wu J., Wen W. Point-of-care testing detection methods for COVID-19. Lab Chip. 2021;21:1634–1660. doi: 10.1039/D0LC01156H. - DOI - PubMed

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