Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests

Abstract DNA synthesis in vitro has enabled the rapid production of reference standards. These are used as controls, and allow measurement and improvement of the accuracy and quality of diagnostic tests. Current reference standards typically represent target genetic material, and act only as positiv...

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Autores principales:	Bindu Swapna Madala, Andre L. M. Reis, Ira W. Deveson, William Rawlinson, Tim R. Mercer
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/eb89f5f1c1f448aba3c47d8c7f9960b9
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spelling	oai:doaj.org-article:eb89f5f1c1f448aba3c47d8c7f9960b92021-12-02T13:57:26ZChimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests10.1038/s41598-021-81760-02045-2322https://doaj.org/article/eb89f5f1c1f448aba3c47d8c7f9960b92021-01-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-81760-0https://doaj.org/toc/2045-2322Abstract DNA synthesis in vitro has enabled the rapid production of reference standards. These are used as controls, and allow measurement and improvement of the accuracy and quality of diagnostic tests. Current reference standards typically represent target genetic material, and act only as positive controls to assess test sensitivity. However, negative controls are also required to evaluate test specificity. Using a pair of chimeric A/B RNA standards, this allowed incorporation of positive and negative controls into diagnostic testing for the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). The chimeric standards constituted target regions for RT-PCR primer/probe sets that are joined in tandem across two separate synthetic molecules. Accordingly, a target region that is present in standard A provides a positive control, whilst being absent in standard B, thereby providing a negative control. This design enables cross-validation of positive and negative controls between the paired standards in the same reaction, with identical conditions. This enables control and test failures to be distinguished, increasing confidence in the accuracy of results. The chimeric A/B standards were assessed using the US Centres for Disease Control real-time RT-PCR protocol, and showed results congruent with other commercial controls in detecting SARS-CoV-2 in patient samples. This chimeric reference standard design approach offers extensive flexibility, allowing representation of diverse genetic features and distantly related sequences, even from different organisms.Bindu Swapna MadalaAndre L. M. ReisIra W. DevesonWilliam RawlinsonTim R. MercerNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-8 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Bindu Swapna Madala Andre L. M. Reis Ira W. Deveson William Rawlinson Tim R. Mercer Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests
description	Abstract DNA synthesis in vitro has enabled the rapid production of reference standards. These are used as controls, and allow measurement and improvement of the accuracy and quality of diagnostic tests. Current reference standards typically represent target genetic material, and act only as positive controls to assess test sensitivity. However, negative controls are also required to evaluate test specificity. Using a pair of chimeric A/B RNA standards, this allowed incorporation of positive and negative controls into diagnostic testing for the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). The chimeric standards constituted target regions for RT-PCR primer/probe sets that are joined in tandem across two separate synthetic molecules. Accordingly, a target region that is present in standard A provides a positive control, whilst being absent in standard B, thereby providing a negative control. This design enables cross-validation of positive and negative controls between the paired standards in the same reaction, with identical conditions. This enables control and test failures to be distinguished, increasing confidence in the accuracy of results. The chimeric A/B standards were assessed using the US Centres for Disease Control real-time RT-PCR protocol, and showed results congruent with other commercial controls in detecting SARS-CoV-2 in patient samples. This chimeric reference standard design approach offers extensive flexibility, allowing representation of diverse genetic features and distantly related sequences, even from different organisms.
format	article
author	Bindu Swapna Madala Andre L. M. Reis Ira W. Deveson William Rawlinson Tim R. Mercer
author_facet	Bindu Swapna Madala Andre L. M. Reis Ira W. Deveson William Rawlinson Tim R. Mercer
author_sort	Bindu Swapna Madala
title	Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests
title_short	Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests
title_full	Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests
title_fullStr	Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests
title_full_unstemmed	Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests
title_sort	chimeric synthetic reference standards enable cross-validation of positive and negative controls in sars-cov-2 molecular tests
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/eb89f5f1c1f448aba3c47d8c7f9960b9
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Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests

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