Ab189945

Tumor tissues from patients were fixed in 37% formalin and embedded in paraffin. After retrieval of antigens, sections were stained with primary antibodies against TGF-β1 (Abcam, ab215715; 1:500), SDHD (Abcam, ab189945; 1:200), and phospho-Stat1 (Cell Signaling, Cat No. 9167; 1:500) at 4 °C overnight. Immunohistochemical staining was performed using a DAB Horseradish Peroxidase Color Development Kit (Beyotime, Shanghai, China) according to the supplied protocol. In brief, tissue slides were stained with HRP-conjugated secondary antibodies (Thermo, Cat No. G-21234, 1:1000) for 1 h at room temperature and stained with hematoxylin (Solarbio, Beijing, China). For immunofluorescent staining, tissue slides were incubated with secondary antibody followed by incubation with DAPI (Solarbio, Beijing, China). The intensity of the immunostaining was analyzed by ImageJ 9.0 software. In each sample, 25 regions were pictured randomly, and the mean expression value of 25 regions was determined as the relative expression value of protein in this sample. In each group, including the CR and CS groups, five tumor tissues were collected and analyzed for difference analysis.