Phenol red as electrochemical indicator for highly sensitive quantification of SARS-CoV-2 by loop-mediated isothermal amplification detection
Copyright © 2023. Published by Elsevier B.V..
The current COVID-19 pandemic has made patent the need for rapid and cost-effective diagnostic tests, crucial for future infectious outbreaks. Loop-mediated isothermal amplification (LAMP) is a promising and decentralized alternative to qPCR. In this work we have developed a sensitive, fast, and simple innovative methodology for quantification of SARS-CoV-2 RNA copies, combining reverse-transcription LAMP with electrochemical detection. This is based on the oxidation of phenol red (PR), a visual and electroactive LAMP indicator, which oxidation peak potential (Ep) changes with the progress of the LAMP reaction. Using that Ep shift as analytical signal, a calibration curve was obtained for fragment N1 copies of SARS-CoV2 (which provided better results than N or S fragments), with a potential shift of 16.2 mV per order of magnitude, and a practical limit of detection of 21 copies·μL-1. Moreover, the precision of Ep is excellent (RSD < 2%): 557 ± 5 mV for negative and 602 ± 7 mV for positive (2148 N fragment RNA copies·µL-1·-1) LAMP controls. This methodology has been applied to the analysis of nasopharyngeal swab samples, resulting in total concordance with clinical RT-qPCR results. Advances towards fully decentralization have been achieved by designing and fabricating a small portable heater for isothermal procedures, obtaining comparable results to those from a commercial benchtop thermal cycler.
| Media Type: |
Electronic Article |
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| Year of Publication: |
2024 2023 |
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| Publication: |
2024 |
| Contained In: |
To Main Record - volume:266 |
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| Contained In: |
Talanta - 266(2023), Pt 1 vom: 01. Jan., Seite 124963 |
| Language: |
English |
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| Contributors: |
Rioboó-Legaspi, P [Author] |
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| Links: |
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| Keywords: |
Electrochemical detection |
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| Notes: |
Date Completed 20.09.2023 Date Revised 20.09.2023 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1016/j.talanta.2023.124963 |
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| funding: |
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| Supporting institution / Project title: |
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| 520 | |a The current COVID-19 pandemic has made patent the need for rapid and cost-effective diagnostic tests, crucial for future infectious outbreaks. Loop-mediated isothermal amplification (LAMP) is a promising and decentralized alternative to qPCR. In this work we have developed a sensitive, fast, and simple innovative methodology for quantification of SARS-CoV-2 RNA copies, combining reverse-transcription LAMP with electrochemical detection. This is based on the oxidation of phenol red (PR), a visual and electroactive LAMP indicator, which oxidation peak potential (Ep) changes with the progress of the LAMP reaction. Using that Ep shift as analytical signal, a calibration curve was obtained for fragment N1 copies of SARS-CoV2 (which provided better results than N or S fragments), with a potential shift of 16.2 mV per order of magnitude, and a practical limit of detection of 21 copies·μL-1. Moreover, the precision of Ep is excellent (RSD < 2%): 557 ± 5 mV for negative and 602 ± 7 mV for positive (2148 N fragment RNA copies·µL-1·-1) LAMP controls. This methodology has been applied to the analysis of nasopharyngeal swab samples, resulting in total concordance with clinical RT-qPCR results. Advances towards fully decentralization have been achieved by designing and fabricating a small portable heater for isothermal procedures, obtaining comparable results to those from a commercial benchtop thermal cycler | ||
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