tetano
Editor, Senior Moderator
Dis Markers
. 2021 Mar 2;2021:8890221.
doi: 10.1155/2021/8890221. eCollection 2021.
Validation of a One-Step Reverse Transcription-Droplet Digital PCR (RT-ddPCR) Approach to Detect and Quantify SARS-CoV-2 RNA in Nasopharyngeal Swabs
Catia Mio[SUP] 1 [/SUP], Adriana Cif?[SUP] 1 [/SUP], Stefania Marzinotto[SUP] 2 [/SUP], Barbara Marcon[SUP] 2 [/SUP], Corrado Pipan[SUP] 1 2 [/SUP], Giuseppe Damante[SUP] 1 2 [/SUP], Francesco Curcio[SUP] 1 2 [/SUP]
Affiliations
Abstract
Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has rapidly spread worldwide from the beginning of 2020. Quantitative reverse transcription-PCR (RT-qPCR) is, to this day, the preferred methodology for viral RNA detection, even if not without problems. To overcome some of the limitations still existing for the detection and quantification of nucleic acids in various applications, the use of one-step reverse transcription-droplet digital PCR (RT-ddPCR) has been established. The purpose of this study was, then, to evaluate the efficacy of ddPCR for the detection of SARS-CoV-2 RNA in nasopharyngeal swabs, optimizing the detection of low-viral load-burdened samples.
Methods: The RT-ddPCR workflow was validated for sensitivity, specificity, linearity, reproducibility, and precision using samples from 90 COVID-19-infected patients referred to the Department of Laboratory Medicine of the University Hospital of Udine (Italy).
Results: The present study shows that RT-ddPCR allows the detection of as low as 10.3 copies of a SARS-COV-2 E-gene per sample with a higher level of accuracy and precision, especially at low concentration.
Conclusion: During the postpeak phase of the SARS-CoV-2 pandemic, it is essential to rely on a highly robust molecular biology method to identify infected subjects, whether they have symptoms or not, in order to prepare appropriate containment measures.
. 2021 Mar 2;2021:8890221.
doi: 10.1155/2021/8890221. eCollection 2021.
Validation of a One-Step Reverse Transcription-Droplet Digital PCR (RT-ddPCR) Approach to Detect and Quantify SARS-CoV-2 RNA in Nasopharyngeal Swabs
Catia Mio[SUP] 1 [/SUP], Adriana Cif?[SUP] 1 [/SUP], Stefania Marzinotto[SUP] 2 [/SUP], Barbara Marcon[SUP] 2 [/SUP], Corrado Pipan[SUP] 1 2 [/SUP], Giuseppe Damante[SUP] 1 2 [/SUP], Francesco Curcio[SUP] 1 2 [/SUP]
Affiliations
- PMID: 33747257
- PMCID: PMC7934789
- DOI: 10.1155/2021/8890221
Abstract
Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has rapidly spread worldwide from the beginning of 2020. Quantitative reverse transcription-PCR (RT-qPCR) is, to this day, the preferred methodology for viral RNA detection, even if not without problems. To overcome some of the limitations still existing for the detection and quantification of nucleic acids in various applications, the use of one-step reverse transcription-droplet digital PCR (RT-ddPCR) has been established. The purpose of this study was, then, to evaluate the efficacy of ddPCR for the detection of SARS-CoV-2 RNA in nasopharyngeal swabs, optimizing the detection of low-viral load-burdened samples.
Methods: The RT-ddPCR workflow was validated for sensitivity, specificity, linearity, reproducibility, and precision using samples from 90 COVID-19-infected patients referred to the Department of Laboratory Medicine of the University Hospital of Udine (Italy).
Results: The present study shows that RT-ddPCR allows the detection of as low as 10.3 copies of a SARS-COV-2 E-gene per sample with a higher level of accuracy and precision, especially at low concentration.
Conclusion: During the postpeak phase of the SARS-CoV-2 pandemic, it is essential to rely on a highly robust molecular biology method to identify infected subjects, whether they have symptoms or not, in order to prepare appropriate containment measures.