Antigen retrieval solution

Immunohistochemical staining was conducted as described previously^{4 (link)}. After treated with 3% hydrogen peroxide to eliminate endogenous peroxidase activity and with Antigen Retrieval Solution (Wuhan Boster Biological Technology Ltd. China) to digest the antigenic sites, the sections were incubated overnight with anti-LOXL2 (1:400, Santa Cruz, sc-48724, USA), anti-TNF-a (1:1000, ab6671, Abcam, USA), or anti-Collagen II (1:100, KG22256, KeyGEN Biotech) at 4 °C. Next, the specimens were incubated with biotin-labeled IgG (BeiJing ZhongShan Golden Bridge Biotechnology Co., China) for 30 min at 37 °C and an avidin-peroxidase complex for 30 min at 37 °C. Then, the specimens were incubated with a peroxidase/diaminobenzidine (DAB) yellow kit (Wuhan Boster Biological Technology Ltd., China) to stain antibody, and counterstained with hematoxylin, dehydrated in an ethanol series, cleared in xylene and cover slipped. Images were acquired as above for the HE staining sections. A color difference was used to delineate positive and negative areas, which were measured with Image-Pro Plus software.