Histological Analysis of Mouse Pancreas

Mouse pancreas tissues were fixed overnight in 4% paraformaldehyde (Sigma-Aldrich, Cat# 158127). Haematoxylin and eosin (H & E) staining and immunostaining were performed to analyse representative cross-sections of the pancreatic head, body and tail. Immunostaining was conducted according to standard protocols [30 (link)]. In brief, endogenous peroxide activity was blocked with 3% peroxidase, and the slides were then incubated with 5% BSA to prevent nonspecific binding. The slides were incubated with primary antibodies at an optimal dilution (clusterin: Santa Cruz Biotechnology, Cat# sc-8354, 1:100; cytokeratin 19: Abcam, Cat# ab15463, 1:100; chymotrypsin: Abcam, Cat# ab35694, 1:100; and amylase: Santa Cruz Biotechnology, Cat# sc-46657, 1:100) and appropriate secondary antibodies. The horseradish peroxidase polymer detection system (PV-9001; GBI Labs, Mukilteo, WA, USA) was employed to detect antibody staining. Five to ten randomly selected 100X H&E images were quantified by a pathologist using standard pathological criteria [31 (link), 32 (link)].