Immunohistochemistry

OGDH expression was detected following standard immunohistochemical protocol as reported elsewhere (19). Briefly, paraffin-embedded slides (5 µM) were baked at 60 °C, then dewaxed by xylene and rehydrated by fractionated ethanol. Antigen was extracted, and incubated with 3% H₂O₂ for 10 min. The slides were then sealed with bovine serum for 1 hour, and anti-OGDH antibody (cat. no. ab137773, Abcam, USA; 1:50 dilution in 1% bovine serum albumin) were then added and incubated at room temperature for 3 hours. The slides were covered with the secondary antibody and placed in a humid chamber for 1 hour, and DAB was added to reveal the staining intensity. Sections were photographed using an Olympus IX81 microscope (Olympus, Inc., Japan).