An Open One-Step RT-qPCR for SARS-CoV-2 detection.
The COVID-19 pandemic has resulted in millions of deaths globally, and while several diagnostic systems were proposed, real-time reverse transcription polymerase chain reaction (RT-PCR) remains the gold standard. However, diagnostic reagents, including enzymes used in RT-PCR, are subject to centrali...
| Main Authors: | , , , , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2024-01-01
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| Series: | PLoS ONE |
| Online Access: | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0297081&type=printable |
| _version_ | 1797326256809705472 |
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| author | Ariel Cerda Maira Rivera Grace Armijo Catalina Ibarra-Henriquez Javiera Reyes Paula Blázquez-Sánchez Javiera Avilés Aníbal Arce Aldo Seguel Alexander J Brown Yesseny Vásquez Marcelo Cortez-San Martín Francisco A Cubillos Patricia García Marcela Ferres César A Ramírez-Sarmiento Fernán Federici Rodrigo A Gutiérrez |
| author_facet | Ariel Cerda Maira Rivera Grace Armijo Catalina Ibarra-Henriquez Javiera Reyes Paula Blázquez-Sánchez Javiera Avilés Aníbal Arce Aldo Seguel Alexander J Brown Yesseny Vásquez Marcelo Cortez-San Martín Francisco A Cubillos Patricia García Marcela Ferres César A Ramírez-Sarmiento Fernán Federici Rodrigo A Gutiérrez |
| author_sort | Ariel Cerda |
| collection | DOAJ |
| description | The COVID-19 pandemic has resulted in millions of deaths globally, and while several diagnostic systems were proposed, real-time reverse transcription polymerase chain reaction (RT-PCR) remains the gold standard. However, diagnostic reagents, including enzymes used in RT-PCR, are subject to centralized production models and intellectual property restrictions, which present a challenge for less developed countries. With the aim of generating a standardized One-Step open RT-qPCR protocol to detect SARS-CoV-2 RNA in clinical samples, we purified and tested recombinant enzymes and a non-proprietary buffer. The protocol utilized M-MLV RT and Taq DNA pol enzymes to perform a Taqman probe-based assay. Synthetic RNA samples were used to validate the One-Step RT-qPCR components, demonstrating sensitivity comparable to a commercial kit routinely employed in clinical settings for patient diagnosis. Further evaluation on 40 clinical samples (20 positive and 20 negative) confirmed its comparable diagnostic accuracy. This study represents a proof of concept for an open approach to developing diagnostic kits for viral infections and diseases, which could provide a cost-effective and accessible solution for less developed countries. |
| first_indexed | 2024-03-08T06:20:48Z |
| format | Article |
| id | doaj.art-d5f833a8615248abb09f8687711811b3 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-03-08T06:20:48Z |
| publishDate | 2024-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-d5f833a8615248abb09f8687711811b32024-02-04T05:31:36ZengPublic Library of Science (PLoS)PLoS ONE1932-62032024-01-01191e029708110.1371/journal.pone.0297081An Open One-Step RT-qPCR for SARS-CoV-2 detection.Ariel CerdaMaira RiveraGrace ArmijoCatalina Ibarra-HenriquezJaviera ReyesPaula Blázquez-SánchezJaviera AvilésAníbal ArceAldo SeguelAlexander J BrownYesseny VásquezMarcelo Cortez-San MartínFrancisco A CubillosPatricia GarcíaMarcela FerresCésar A Ramírez-SarmientoFernán FedericiRodrigo A GutiérrezThe COVID-19 pandemic has resulted in millions of deaths globally, and while several diagnostic systems were proposed, real-time reverse transcription polymerase chain reaction (RT-PCR) remains the gold standard. However, diagnostic reagents, including enzymes used in RT-PCR, are subject to centralized production models and intellectual property restrictions, which present a challenge for less developed countries. With the aim of generating a standardized One-Step open RT-qPCR protocol to detect SARS-CoV-2 RNA in clinical samples, we purified and tested recombinant enzymes and a non-proprietary buffer. The protocol utilized M-MLV RT and Taq DNA pol enzymes to perform a Taqman probe-based assay. Synthetic RNA samples were used to validate the One-Step RT-qPCR components, demonstrating sensitivity comparable to a commercial kit routinely employed in clinical settings for patient diagnosis. Further evaluation on 40 clinical samples (20 positive and 20 negative) confirmed its comparable diagnostic accuracy. This study represents a proof of concept for an open approach to developing diagnostic kits for viral infections and diseases, which could provide a cost-effective and accessible solution for less developed countries.https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0297081&type=printable |
| spellingShingle | Ariel Cerda Maira Rivera Grace Armijo Catalina Ibarra-Henriquez Javiera Reyes Paula Blázquez-Sánchez Javiera Avilés Aníbal Arce Aldo Seguel Alexander J Brown Yesseny Vásquez Marcelo Cortez-San Martín Francisco A Cubillos Patricia García Marcela Ferres César A Ramírez-Sarmiento Fernán Federici Rodrigo A Gutiérrez An Open One-Step RT-qPCR for SARS-CoV-2 detection. PLoS ONE |
| title | An Open One-Step RT-qPCR for SARS-CoV-2 detection. |
| title_full | An Open One-Step RT-qPCR for SARS-CoV-2 detection. |
| title_fullStr | An Open One-Step RT-qPCR for SARS-CoV-2 detection. |
| title_full_unstemmed | An Open One-Step RT-qPCR for SARS-CoV-2 detection. |
| title_short | An Open One-Step RT-qPCR for SARS-CoV-2 detection. |
| title_sort | open one step rt qpcr for sars cov 2 detection |
| url | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0297081&type=printable |
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