Immunohistochemical Analysis of SARS-CoV-2 Entry Factors

Surgical resections were fixed in formalin within minutes of surgery.45 (link) After processing, the resected small airway (<2 mm internal diameter) tissue blocks were separately embedded in paraffin wax for research analyses. The tissue was sectioned at 3.5 µm and processed with standard immunohistochemical staining procedures.45 (link) Immunostaining was completed by using rabbit polyclonal anti-ACE2 antibody (Catalog No. Ab15348, Abcam, 1:800), Furin rabbit polyclonal antibody (bs-13228R; Bioss antibodies; 1:200) and TMPRSS2 rabbit polyclonal antibody (bs-6285R, Bioss antibodies, 1:250). Antibody binding was visualised by a substrate 3,3ʹ-diaminobenzidine (DAB) reaction producing a brown colour indicating positive staining. Nuclear counterstain was achieved with gills hematoxylin. All slides were coded and randomised to blind the analyst.

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Brake S.J., Eapen M.S., McAlinden K.D., Markos J., Haug G., Larby J., Chia C., Hardikar A., Singhera G.K., Hackett T.L., Lu W, & Sohal S.S. (2022). SARS-CoV-2 (COVID-19) Adhesion Site Protein Upregulation in Small Airways, Type 2 Pneumocytes, and Alveolar Macrophages of Smokers and COPD – Possible Implications for Interstitial Fibrosis. International Journal of Chronic Obstructive Pulmonary Disease, 17, 101-115.

Publication 2022

bs-13228R bs-6285R

Ace2 Anti antibody Antibodies Antibody Blind Formalin Furin Gills Hematoxylin Paraffin wax Rabbit Surgery Tissue Tmprss2

Corresponding Organization : University of Tasmania

Other organizations : Launceston General Hospital, Royal Hobart Hospital, University of British Columbia, St. Paul's Hospital

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Variable analysis

independent variables

Tissue processing: Surgical resections were fixed in formalin within minutes of surgery, and the resected small airway (<2 mm internal diameter) tissue blocks were separately embedded in paraffin wax for research analyses.

dependent variables

Immunostaining results: Antibody binding was visualised by a substrate 3,3ʹ-diaminobenzidine (DAB) reaction producing a brown colour indicating positive staining.

control variables

Tissue sectioning: The tissue was sectioned at 3.5 µm and processed with standard immunohistochemical staining procedures.
Blinding: All slides were coded and randomised to blind the analyst.

controls

Positive control: Not explicitly mentioned.
Negative control: Not explicitly mentioned.

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