MicroRNA ISH Buffer and Controls kit (Exiqon A/S, Vedbaek, Denmark) were used for ISH in the present study, according to the manufacturer's protocol. The ISH detection of miR-1, miR-10b and miR-214 in tumor tissues were performed as described previously (22 (link)). Briefly, formalin-fixed paraffin-embedded PDAC tissues were cut into 4-µm sections and deparaffinized (23 (link)). The probes for miR-1, miR-10b and miR-214 used in the present study were complementary to human mature miR-1, miR-10b and miR-214, respectively. For signal detection, anti-digoxigenin-alkaline phosphatase Fab fragments (1:800; Roche Applied Science, Penzburg, Germany; Roche Diagnostics Gmbh, Mannheim, Germany) were used as the primary antibody, and the slides were incubated with nitro blue tetrazolium/5-bromo-4-chloroindol-3-yl-phosphate solution (Roche Applied Science; Roche Diagnostics GmbH). Counterstaining was performed by Nuclear Fast Red (Chroma ATE Inc., Stuttgart, Germany).