Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue
Background: There is increasing evidence that identification of SARS-CoV-2 virions by transmission electron microscopy could be misleading due to the similar morphology of virions and ubiquitous cell structures. This study thus aimed to establish methods for indisputable proof of the presence of SAR...
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| Format: | Article |
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MDPI AG
2021-09-01
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| Series: | Viruses |
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| Online Access: | https://www.mdpi.com/1999-4915/13/9/1816 |
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| author | Andreja Erman Karmen Wechtersbach Daniel Velkavrh Jerica Pleško Maja Frelih Nika Kojc |
| author_facet | Andreja Erman Karmen Wechtersbach Daniel Velkavrh Jerica Pleško Maja Frelih Nika Kojc |
| author_sort | Andreja Erman |
| collection | DOAJ |
| description | Background: There is increasing evidence that identification of SARS-CoV-2 virions by transmission electron microscopy could be misleading due to the similar morphology of virions and ubiquitous cell structures. This study thus aimed to establish methods for indisputable proof of the presence of SARS-CoV-2 virions in the observed tissue. Methods: We developed a variant of the correlative microscopy approach for SARS-CoV-2 protein identification using immunohistochemical labelling of SARS-CoV-2 proteins on light and electron microscopy levels. We also performed immunogold labelling of SARS-CoV-2 virions. Results: Immunohistochemistry (IHC) of SARS-CoV-2 nucleocapsid proteins and subsequent correlative microscopy undoubtedly proved the presence of SARS-CoV-2 virions in the analysed human nasopharyngeal tissue. The presence of SARS-CoV-2 virions was also confirmed by immunogold labelling for the first time. Conclusions: Immunoelectron microscopy is the most reliable method for distinguishing intracellular viral particles from normal cell structures of similar morphology and size as virions. Furthermore, we developed a variant of correlative microscopy that allows pathologists to check the results of IHC performed first on routinely used paraffin-embedded samples, followed by semithin, and finally by ultrathin sections. Both methodological approaches indisputably proved the presence of SARS-CoV-2 virions in cells. |
| first_indexed | 2024-03-10T07:08:51Z |
| format | Article |
| id | doaj.art-9de6eee2e0964187986d2d9ef4e2e445 |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-10T07:08:51Z |
| publishDate | 2021-09-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-9de6eee2e0964187986d2d9ef4e2e4452023-11-22T15:38:26ZengMDPI AGViruses1999-49152021-09-01139181610.3390/v13091816Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected TissueAndreja Erman0Karmen Wechtersbach1Daniel Velkavrh2Jerica Pleško3Maja Frelih4Nika Kojc5Institute of Cell Biology, Faculty of Medicine, University of Ljubljana, 1000 Ljubljana, SloveniaInstitute of Pathology, Faculty of Medicine, University of Ljubljana, 1000 Ljubljana, SloveniaInstitute of Pathology, Faculty of Medicine, University of Ljubljana, 1000 Ljubljana, SloveniaInstitute of Pathology, Faculty of Medicine, University of Ljubljana, 1000 Ljubljana, SloveniaInstitute of Pathology, Faculty of Medicine, University of Ljubljana, 1000 Ljubljana, SloveniaInstitute of Pathology, Faculty of Medicine, University of Ljubljana, 1000 Ljubljana, SloveniaBackground: There is increasing evidence that identification of SARS-CoV-2 virions by transmission electron microscopy could be misleading due to the similar morphology of virions and ubiquitous cell structures. This study thus aimed to establish methods for indisputable proof of the presence of SARS-CoV-2 virions in the observed tissue. Methods: We developed a variant of the correlative microscopy approach for SARS-CoV-2 protein identification using immunohistochemical labelling of SARS-CoV-2 proteins on light and electron microscopy levels. We also performed immunogold labelling of SARS-CoV-2 virions. Results: Immunohistochemistry (IHC) of SARS-CoV-2 nucleocapsid proteins and subsequent correlative microscopy undoubtedly proved the presence of SARS-CoV-2 virions in the analysed human nasopharyngeal tissue. The presence of SARS-CoV-2 virions was also confirmed by immunogold labelling for the first time. Conclusions: Immunoelectron microscopy is the most reliable method for distinguishing intracellular viral particles from normal cell structures of similar morphology and size as virions. Furthermore, we developed a variant of correlative microscopy that allows pathologists to check the results of IHC performed first on routinely used paraffin-embedded samples, followed by semithin, and finally by ultrathin sections. Both methodological approaches indisputably proved the presence of SARS-CoV-2 virions in cells.https://www.mdpi.com/1999-4915/13/9/1816SARS-CoV-2immunolabellinglight microscopyelectron microscopycorrelative microscopy |
| spellingShingle | Andreja Erman Karmen Wechtersbach Daniel Velkavrh Jerica Pleško Maja Frelih Nika Kojc Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue Viruses SARS-CoV-2 immunolabelling light microscopy electron microscopy correlative microscopy |
| title | Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue |
| title_full | Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue |
| title_fullStr | Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue |
| title_full_unstemmed | Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue |
| title_short | Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue |
| title_sort | just seeing is not enough for believing immunolabelling as indisputable proof of sars cov 2 virions in infected tissue |
| topic | SARS-CoV-2 immunolabelling light microscopy electron microscopy correlative microscopy |
| url | https://www.mdpi.com/1999-4915/13/9/1816 |
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