Immunohistochemical Staining of Ki67

Xenograft tumors were collected, and paraffin-embedded sections were prepared. For immunohistochemistry staining, tumor sections were deparaffinized in xylene and rehydrated in graded alcohols. Then, sections were treated with 3% hydrogen peroxide for 15 mins and incubated with 10% goat serum for 30 mins to decrease nonspecific binding. After washing three times in PBS, the sections were incubated with anti-Ki67 (1:500, ZSGB-BIO, China, ZM-0166) at 4°C overnight, followed by incubation with peroxidase labeled secondary antibody at room temperature for 30 mins. The nuclei were counterstained with hematoxylin for 5 min, dehydrated, and mounted for microscopic examination.