Anti cd44

Protein samples were extracted from carotid arteries with a lysis buffer. The protein concentration was determined by the BCA method (Beyotime Institute of Biotechnology, Jiangsu, China). All samples were separated with different concentrations of SDS–PAGE according to different molecular weights, and transferred to an Immun-Blot polyvinylidene difluoride membrane (Bio-Rad, Hercules, CA, USA), membranes were blocked in 5% BSA (Solarbio, Beijing, China) in 1xTBST and were then incubated with anti-β-Actin (CTS, 8H10D10), anti-Ptprc (20103-1-AP; Proteintech, Rosemont, IL, USA), anti-Itgb2 (10554-1-AP; Proteintech, Rosemont, IL, USA), anti-Itgax (GB11059; Servicebio, Wuhan, China), anti-Ctss (Santa Cruz, sc-271619), anti-Ly86 (Santa Cruz, sc-390613), anti-CD44 (GB112054; Servicebio, Wuhan, China), anti-Aif1 (Santa Cruz, sc-32725) primary antibodies with working concentration about 1 μg/mL, and a horseradish peroxidase–conjugated secondary antibody, respectively. The bands were visualized using the Super Western Sensitivity Chemiluminescence Detection System (Thermo Fisher, Waltham, MA, USA). Autoradiographs were quantified by densitometry (NIH Image J).

IHC was performed as previously described (Zhang and Gu, 2022 (link)). Briefly, 5-µm thick formalin-fixed paraffin-embedded carotid tissue of mice sections were stained with anti-Cd44 (Servicebio), anti-Il1b (Servicebio), anti-Ptgs2 (Servicebio), and anti-Cybb (Servicebio) antibodies according to the manufacturer’s instructions. All positive cells were counted from three sections of each artery sample and evaluated by an investigator who was blinded to the identities of the treatment protocols at ×100 magnification.