Pias3

Paraffin-embedded xenografts were sliced into 7 μm sections and subjected to HE morphological staining and immunohistochemical staining by the methods described elsewhere.12 (link) Because ARHI has been known as OC suppressor24 (link) and STAT3 signaling as the main resveratrol molecular target,25 their statuses in the orthotopic tumor tissues with and without resveratrol treatment were analyzed. The antibodies used were mouse anti-ARHI (1:100; Santa Cruz, CA, USA), PIAS3 (1:120; Santa Cruz, CA, USA) and phosphorylated STAT3/p-STAT3 (1:300; ProteinTech, Chicago, IL, USA), respectively. Binding of the primary antibody was detected by a peroxidase reaction using 3,3-diaminobenzidine tetrahydrochloride (DAB) as the substrate (Vector Laboratories, Burlingame, CA, USA). According to the labeling intensity, the staining results were evaluated by two independent researchers and scored as negative (-) if no immunolabeling was observed in target cells, weakly positive (+) if the labeling was faint, moderately positive (++), and strongly positive (+++) when the labeling was stronger or distinctly stronger than (++). Tissue sections without primary antibody incubation were used as background controls for IHC staining and β-actin was used as a quantitative control for Western blotting.