Immunofluorescence Analysis of Pancreatic Tissue

Pancreatic tissue sections from βUCP2Tg mice and in vitro-cultured human islets were immunostained with anti-insulin (Abcam), anti-UCP2 (Santa Cruz), anti-AldB (Proteintech), or anti-BrdU (Dako, Tokyo, Japan) antibodies. Images were acquired as described elsewhere (Shirakawa et al., 2013 (link)). For immunofluorescence cytochemistry (IFCC), 1 × 10⁴ MIN6-M9 cells were seeded on poly-L-lysine-coated chamber slides (Nunk, #154534PK) and infected with Ad-LacZ or Ad-Ucp2 at an MOI of 500 for 48 h. Cells were fixed with 4% PFA and were permeabilized with 0.01% Triton X-100 in PBS. Anti-HNF4α antibody was purchased from Abcam (#ab201460). Images were acquired using an FV1000-D confocal laser scanning microscope (Olympus, Japan). Super resolution STED microscopic images were acquired using TCS SP8 STED (Leica Microsystems, Germany).