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Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens
The rapid emergence of SARS-CoV-2, the causative agent of COVID-19, and its dissemination globally has caused an unprecedented strain on public health. Animal models are urgently being developed for SARS-CoV-2 to aid rational design of vaccines and therapeutics. Immunohistochemistry and in situ hybr...
Autores principales: | , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7736337/ https://www.ncbi.nlm.nih.gov/pubmed/33318594 http://dx.doi.org/10.1038/s41598-020-78949-0 |
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author | Lean, Fabian Z. X. Lamers, Mart M. Smith, Samuel P. Shipley, Rebecca Schipper, Debby Temperton, Nigel Haagmans, Bart L. Banyard, Ashley C. Bewley, Kevin R. Carroll, Miles W. Brookes, Sharon M. Brown, Ian Nuñez, Alejandro |
author_facet | Lean, Fabian Z. X. Lamers, Mart M. Smith, Samuel P. Shipley, Rebecca Schipper, Debby Temperton, Nigel Haagmans, Bart L. Banyard, Ashley C. Bewley, Kevin R. Carroll, Miles W. Brookes, Sharon M. Brown, Ian Nuñez, Alejandro |
author_sort | Lean, Fabian Z. X. |
collection | PubMed |
description | The rapid emergence of SARS-CoV-2, the causative agent of COVID-19, and its dissemination globally has caused an unprecedented strain on public health. Animal models are urgently being developed for SARS-CoV-2 to aid rational design of vaccines and therapeutics. Immunohistochemistry and in situ hybridisation techniques that facilitate reliable and reproducible detection of SARS-CoV and SARS-CoV-2 viral products in formalin-fixed paraffin-embedded (FFPE) specimens would be of great utility. A selection of commercial antibodies generated against SARS-CoV spike protein and nucleoprotein, double stranded RNA, and RNA probe for spike genes were evaluated for the ability to detect FFPE infected cells. We also tested both heat- and enzymatic-mediated virus antigen retrieval methods to determine the optimal virus antigen recovery as well as identifying alternative retrieval methods to enable flexibility of IHC methods. In addition to using native virus infected cells as positive control material, the evaluation of non-infected cells expressing coronavirus (SARS, MERS) spike as a biosecure alternative to assays involving live virus was undertaken. Optimized protocols were successfully applied to experimental animal-derived tissues. The diverse techniques for virus detection and control material generation demonstrated in this study can be applied to investigations of coronavirus pathogenesis and therapeutic research in animal models. |
format | Online Article Text |
id | pubmed-7736337 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-77363372020-12-15 Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens Lean, Fabian Z. X. Lamers, Mart M. Smith, Samuel P. Shipley, Rebecca Schipper, Debby Temperton, Nigel Haagmans, Bart L. Banyard, Ashley C. Bewley, Kevin R. Carroll, Miles W. Brookes, Sharon M. Brown, Ian Nuñez, Alejandro Sci Rep Article The rapid emergence of SARS-CoV-2, the causative agent of COVID-19, and its dissemination globally has caused an unprecedented strain on public health. Animal models are urgently being developed for SARS-CoV-2 to aid rational design of vaccines and therapeutics. Immunohistochemistry and in situ hybridisation techniques that facilitate reliable and reproducible detection of SARS-CoV and SARS-CoV-2 viral products in formalin-fixed paraffin-embedded (FFPE) specimens would be of great utility. A selection of commercial antibodies generated against SARS-CoV spike protein and nucleoprotein, double stranded RNA, and RNA probe for spike genes were evaluated for the ability to detect FFPE infected cells. We also tested both heat- and enzymatic-mediated virus antigen retrieval methods to determine the optimal virus antigen recovery as well as identifying alternative retrieval methods to enable flexibility of IHC methods. In addition to using native virus infected cells as positive control material, the evaluation of non-infected cells expressing coronavirus (SARS, MERS) spike as a biosecure alternative to assays involving live virus was undertaken. Optimized protocols were successfully applied to experimental animal-derived tissues. The diverse techniques for virus detection and control material generation demonstrated in this study can be applied to investigations of coronavirus pathogenesis and therapeutic research in animal models. Nature Publishing Group UK 2020-12-14 /pmc/articles/PMC7736337/ /pubmed/33318594 http://dx.doi.org/10.1038/s41598-020-78949-0 Text en © The Author(s) 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. |
spellingShingle | Article Lean, Fabian Z. X. Lamers, Mart M. Smith, Samuel P. Shipley, Rebecca Schipper, Debby Temperton, Nigel Haagmans, Bart L. Banyard, Ashley C. Bewley, Kevin R. Carroll, Miles W. Brookes, Sharon M. Brown, Ian Nuñez, Alejandro Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens |
title | Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens |
title_full | Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens |
title_fullStr | Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens |
title_full_unstemmed | Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens |
title_short | Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens |
title_sort | development of immunohistochemistry and in situ hybridisation for the detection of sars-cov and sars-cov-2 in formalin-fixed paraffin-embedded specimens |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7736337/ https://www.ncbi.nlm.nih.gov/pubmed/33318594 http://dx.doi.org/10.1038/s41598-020-78949-0 |
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