Recombinant Anti-SOX2 antibody [EPR3131] (ab92494)

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Formalin-fixed, paraffin-embedded blocks of 341 UTUC specimens were retrieved from the Department of Urological Pathology. The expression of SOX2 was evaluated by standard IHC protocols. In the case of giant or multiple tumors, only the largest tumor specimen was processed for IHC analysis. Briefly, 4-mm sections from all formalin-fixed, paraffin-embedded specimens were deparaffinized with xylene and rehydrated in decreasing concentrations of ethanol. Heat-induced antigen retrieval was performed using sodium citrate in a pressure cooker at 120°C for 20 min. After blocking endogenous peroxidases with 3% hydrogen peroxide in methanol, the sections were incubated with 10% normal blocking serum in Tris-buffered saline for 20 min. Based on preliminary evaluations, the sections were incubated with an anti-SOX2 antibody (ab92494; 1:200; Abcam, Hong Kong, China) overnight, followed by incubation with a goat anti-rabbit IgG/HRP polymer (PV-9001, ZSGB-BIO, Beijing, China) for 20 min. The sections were then exposed to a diaminobenzidine tetrahydrochloride solution and counterstained with hematoxylin. The stained tissue sections were reviewed and scored by two independent urological pathologists without any prior knowledge of the clinical data of the cohort (X.Y. and Q.H.), disagreements were reconciled by multi-headed consensus review. Positive SOX2 expression was defined as ≥10% tumor cells with positive nuclear staining.15,17

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