Immunohistochemical Analysis of GC Tissues

GC tissue specimens were cut into 5-μm-thick sections. The sections were blocked with normal goat serum and immunostained with primary antibodies to active YAP, CDX2 and Rab27a overnight at 4 ºC, and then with biotin-labeled secondary antibody goat anti-rabbit or goat anti-mouse (BM3894 or BM3895, 1: 500, Boster). Next, the sections were incubated with 50 μL streptomyces anti-biotin–peroxidase solution at room temperature for 10 min and developed with DAB. Following counterstaining, dehydration, clearing and mounting, the sections were observed with a microscope. Primary antibody was substituted by PBS. Positive cells were brown-yellow and the sum of the integrated optical density (IOD) was analyzed using Image-ProPlus6.0 [20 (link)].