Development of immunohistochemistry and in situ hybridisation for the detection of SARS-CoV and SARS-CoV-2 in formalin-fixed paraffin-embedded specimens

Fabian Z.X. Lean, Mart M. Lamers, Samuel P. Smith, Rebecca Shipley, Debby Schipper, Nigel Temperton, Bart L. Haagmans, Ashley C. Banyard, Kevin R. Bewley, Miles Carroll, Sharon M. Brookes, Ian Brown, Alejandro Nuñez

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13 Citations (Scopus)


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.

Original languageEnglish
Article number21894
JournalScientific Reports
Issue number1
Publication statusPublished - Dec 2020

Bibliographical note

Funding Information:
SARS-CoV HKU-39849 and SARS-CoV-2 BetaCoV/Munich/BavPat1/2020 virus isolates were kindly provided by Professor Malik Peiris, University of Hong Kong and Professor Christian Drosten, Charité Universitätsmedi-zin, respectively. We also appreciate Victorian Infectious Diseases Reference Laboratory (VIDRL) and the Peter Doherty Institute, Australia (Dr Julian Druce) for sharing the virus isolate nCoV/Victoria/2/2020. We would like to thank staff at the histology laboratory at APHA. The study was funded by Department for Environment, Food & Rural Affairs (DEFRA), UK (Project SE0557, EXOM0414, SV3700). Funder has no role in study design, analysis, interpretation or writing of the report.

Publisher Copyright:
© 2020, The Author(s).


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