Immunohistochemistry

Immunohistochemistry was performed on renal cancer specimens as previously described. The evaluation of E2F3 and HIF-2α protein expression was independently and blindly analyzed by two observers (Y. F and YX. Y). According to the previous study, staining intensity was scored 0 (negative), 1 (weak), 2 (moderate), and 3 (strong). For convenience in the Chi-square test, the final score was determined by multiplying the intensity scores with staining extent, and the results were ranged from 0–9. Final scores (intensity score × percentage score) less than 2 were considered as negative staining, 2–3 as weak staining (+), 4–6 as moderate staining (++), and >6 as strong staining (+++)[38].