Horseradish peroxidase conjugated goat anti mouse immunoglobulin g antibody

Tumor xenografts were fixed in formalin solution overnight at room temperature and embedded in paraffin blocks, which were sliced into 4-µm thick sections for immunohistochemical staining. Following deparaffinization, the sections were incubated at 4°C overnight with antibodies of Bcl-2 (cat no. 15071), Bax (cat no. 5023), caspase-3 (cat no. 9662) and survivin (cat no. 2808) (1:500; Cell Signaling Technology, Inc.). The sections were washed 3 times with PBS with 0.1% Tween-20 (PBST), and incubated for 1 h at room temperature with a horseradish peroxidase-conjugated goat anti-mouse immunoglobulin G antibody (1:200; cat no. sc-2031; Santa Cruz Biotechnology, Inc.). The slices were washed with PBST 3 times, and incubated in diaminobenzidine (Sangon Biotech Co., Ltd., Shanghai, China) for 5 min at room temperature and counterstained with hematoxylin (Sangon Biotech Co., Ltd.) for 1 min at room temperature. Images were captured using a light microscope (Olympus Corporation) at a magnification of ×20. Five sections per tumor tissue were examined.