Immunohistochemical (IHC) analysis (Fig. 6c and Supplementary Fig. 6) was performed as previously described2 ,5 (link),10 (link),25 (link)–27 (link) using an Autostainer Link 48 (Dako). The deparaffinized sections were exposed to EnVision FLEX target retrieval solution high pH (Agilent, Cat# K8004) for 20 min at 97 °C for activation, and a mouse anti-SARS-CoV-2 N monoclonal antibody (clone 1035111, R&D Systems, Cat# MAB10474-SP, 1:400) was used as a primary antibody. The sections were sensitized using EnVision FLEX for 15 min and visualized by peroxidase-based enzymatic reaction with 3,3’-diaminobenzidine tetrahydrochloride (Dako, Cat# DM827) as substrate for 5 min. The N protein positivity was evaluated by certificated pathologists as previously described2 ,5 (link),10 (link),25 (link)–27 (link). Images were incorporated as virtual slides by NDP.scan software v3.2.4 (Hamamatsu Photonics). The N-protein positivity was measured as the area using Fiji software v2.2.0 (ImageJ).
Tamura T., Ito J., Uriu K., Zahradnik J., Kida I., Anraku Y., Nasser H., Shofa M., Oda Y., Lytras S., Nao N., Itakura Y., Deguchi S., Suzuki R., Wang L., Begum M.M., Kita S., Yajima H., Sasaki J., Sasaki-Tabata K., Shimizu R., Tsuda M., Kosugi Y., Fujita S., Pan L., Sauter D., Yoshimatsu K., Suzuki S., Asakura H., Nagashima M., Sadamasu K., Yoshimura K., Yamamoto Y., Nagamoto T., Schreiber G., Maenaka K., Hashiguchi T., Ikeda T., Fukuhara T., Saito A., Tanaka S., Matsuno K., Takayama K, & Sato K. (2023). Virological characteristics of the SARS-CoV-2 XBB variant derived from recombination of two Omicron subvariants. Nature Communications, 14, 2800.
Anti antibody Antibody Clone Enzymatic Mouse N proteinPathologistsPeroxidase Sars cov 2 Scan
Corresponding Organization :
Other organizations :
Hokkaido University, The University of Tokyo, Weizmann Institute of Science, Kumamoto University, University of Miyazaki, MRC University of Glasgow Centre for Virus Research, Medical Research Council, Kyoto University, Kyushu University, Tokyo Metropolitan Institute of Public Health, Tokai University, Hiroshima University, Japan Agency for Medical Research and Development, Sapporo University
Antibody used (mouse anti-SARS-CoV-2 N monoclonal antibody, clone 1035111, R&D Systems, Cat# MAB10474-SP)
dependent variables
N protein positivity (measured as the area using Fiji software v2.2.0 (ImageJ))
control variables
Deparaffinized tissue sections
Exposure to EnVision FLEX target retrieval solution high pH (Agilent, Cat# K8004) for 20 min at 97 °C for activation
Sensitization using EnVision FLEX for 15 min
Visualization by peroxidase-based enzymatic reaction with 3,3'-diaminobenzidine tetrahydrochloride (Dako, Cat# DM827) as substrate for 5 min
controls
Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned
Annotations
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