Immunohistochemical Analysis of PPARα

The frozen sections were incubated with 3% (v/v) H₂O₂ for 15 min to quench the endogenous peroxidase activity and blocked with 10% goat serum (ZLI-9021, ORIGINE, Beijing, China) for 1 h at room temperature. The sections were incubated with anti-PPARα (1:100 dilution, PA1-822A, Invitrogen, Waltham, MA, USA) antibodies overnight at 4 °C. The biotinylated secondary antibodies of the enzyme-labeled goat anti-mouse/rabbit IgG polymer (GK600505-B, Gene Tech, Shanghai, China) were added, followed by an incubation at 37 °C for 1 h. The slides were then developed with a DAB chromogenic solution (GK600505, Gene Tech, Shanghai, China) and counterstained with hematoxylin. The images were acquired using an Olympus laser scanning microscope (U-LH100-3), and the intensity of the immunostaining was analyzed by the average of 5 slides with 5 random fields at ×40 per slide using Image J software (NIH, Bethesda, MD, USA) [26 (link)].