RT-qPCR half-reaction optimization for the detection of SARS-CoV-2
| Autor: | Priscila Lamb Wink, Fabiana Volpato, Daiana de Lima-Morales, Rodrigo Minuto Paiva, Julia Biz Willig, Hugo Bock, Fernanda de Paris, Afonso Luís Barth |
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| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: | |
| Zdroj: | Revista da Sociedade Brasileira de Medicina Tropical, Vol 54 (2021) |
| Druh dokumentu: | article |
| ISSN: | 1678-9849 0037-8682 |
| DOI: | 10.1590/0037-8682-0319-2020 |
| Popis: | Abstract INTRODUCTION: The main laboratory test for the diagnosis of coronavirus disease 2019 (COVID-19) is the reverse transcription real-time polymerase chain reaction (RT-qPCR). However, RT-qPCR is expensive because of the number of tests required. This study aimed to evaluate an alternative to the RT-qPCR approach for the detection of sudden acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that is half of the total volume currently recommended by the US Centers for Disease Control and Prevention. METHODS: The analytical limit of detection (LoD) and the reaction efficiency using half volumes of the RT-qPCR assay were evaluated for the N1 and N2 regions using a synthetic control RNA. A panel of 76 SARS-CoV-2-positive and 26 SARS-CoV-2-negative clinical samples was evaluated to establish clinical sensitivity and specificity. RESULTS: The RT-qPCR assay efficiency was 105% for the half and standard reactions considering the N2 target and 84% (standard) and 101% (half) for N1. The RT-qPCR half-reaction LoD for N1 and N2 were 20 and 80 copies/µL, respectively. The clinical sensitivity and specificity were 100%. The half reaction presented a decrease of up to 5.5 cycle thresholds compared with standard RT-qPCR. CONCLUSIONS: The use of the RT-qPCR half-reaction proved feasible and economic for the detection of SARS-CoV-2 RNA. |
| Databáze: | Directory of Open Access Journals |
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