Sabc cy3

The ginsenoside Rk1 (purity > 95%) was isolated from black ginseng as described in our previous work [27] . The APAP (>98.0%, UV-VIS, batch no. A7685-100G) was purchased from Sigma–Aldrich (St Louis, MO, USA). The assay kits for alanine aminotransferase (ALT), aspartate aminotransferase (AST), GSH, superoxide dismutase (SOD), and MDA were provided by Nanjing Jiancheng Biological Research Institute (Nanjing, China). Two-site sandwich enzyme-linked immunosorbent assay (ELISA) kits for the detection of mouse TNF-α and IL-1β were purchased from R&D systems (Minneapolis, MN, USA). Hematoxylin–eosin (H&E) and Hoechst 33258 dye kits were obtained from Beyotime Co., Ltd. (Shanghai, China). Two-site immunohistochemical assay kits, SABC-DyLight 488 and SABC-Cy3, and immunofluorescence staining kits were from BOSTER Biological Technology (Wuhan, China). The antibodies, including rabbit monoclonal anti-iNOS, anti-COX-2, anti-Bax, anti-Bcl-2, anti-cytochrome P450 E1 (CPY2E1), anti-4-HNE, and anti-3-nitrotyrosine (3-NT), were provided by BOSTER Biological Technology (Wuhan, China) or Cell Signaling Technology (Danvers, MA, USA). In situ Cell Death Detection Kit was provided by Roche Applied Science (No. 11684817910; Roche Applied Science, Penzberg, Germany). All other reagents and chemicals, unless indicated, were obtained from Beijing Chemical Factory (Beijing, China).

The mouse aorta, liver, kidney, and pancreas were fixed in 4% paraformaldehyde, embedded in paraffin, sectioned with a paraffin microtome (Leica RM 2245, Wetzlar, Germany) and then HE stained (200×; Solarbio). The mouse aorta was fixed in 4% paraformaldehyde, dehydrated in a sucrose solution gradient, and sectioned with a cryostat (Leica CM 1950). The sections were incubated with anti‐HIF‐1α and anti‐VEGF‐A (1:100; ABclonal, Wuhan, China) at 4°C overnight, and washed with secondary antibody DyLight 488 (1:400; Boster, Wuhan, China) and SABC‐CY3 (1:100; Boster), and then the sections were stained with DAPI (1:1000; Boster) in the dark. A fluorescence microscope (400×; Olympus) and Image‐J software were used to observe and analyze the intensity of the immunofluorescence staining.