Immunohistochemical Detection of HO-1 and SARS-CoV-2

Tissue sections were heat-treated in citric acid buffer and then incubated with 2% H₂O₂ to inactivate the endogenous peroxidase. Following blocking with 3% normal goat serum (NGS) for 1 h, sections were incubated with anti-HO-1 monoclonal antibody (cat no: ADI-SPA-895, Enzo Life Sciences, Farmingdale, NY 11735, USA) at a 1:100 dilution overnight at 4 °C. After incubation with an HRP-conjugated secondary antibody, binding was detected using Vectastain ABC Kit (Vector Laboratories, Burlingame, CA, USA) according to manufacturer’s instructions. The 3,3-diaminobenzidine was used as chromogen and slides were counterstained with haematoxylin and observed under an Olympus (BX50F4) microscope (Centre Valley, PA 18034, USA). For SARS-CoV-2 tissue staining with an anti-SARSCoV-2 (G2) monoclonal antibody [13 (link)] was performed at a 1:300 dilution as previously described [13 (link)]. Routine procedures for Prussian blue (detection of hemosiderin, a ferritin complex) staining were carried out.