Histological Assessment of Pancreas and Liver

Pancreas sections were fixed in 4% paraformaldehyde, embedded in paraffin, cut into 5 μm sections, and rehydrated by xylene and declining grades of ethanol for 5 min. Then, they were washed three times for HE staining. Immunohistochemical and immunofluorescent tests were performed using anti-PDX-1 (Abcam, UK, ab47383) antibody and anti-insulin (Cell Signaling Technology, USA, #4590S) antibody. Frozen livers were sliced, and Oil Red O staining was performed as described earlier [30 (link), 31 (link)] using the Oil Red O Stain Kit (Lipid Stain) (SenBeiJia Biological Technology, China, BP-DL101). According to the instructions of the kit, the Oil Red O solution was added dropwise onto the tissue for a 5-10 min incubation period. Excess staining buffer was removed with 85% propylene glycol. The tissues were washed with distilled water and counterstained with hematoxylin.