Haematoxylin and eosin (H&E) staining for histopathology

HE staining were performed to evaluate the renal pathological changes, glomerular morphological changes and renal fibrosis in the rats kidneys. Kidney tissues from all the rats were fixed with 4% paraformaldehyde at 4˚C for 24 h and embedded in paraffin blocks before they were sectioned at ~3-4 µm thickness. The sections were treated with xylene for 5-10 min, immersed in a mixture of xylene and pure ethanol (1:1) for 5 min and then in 100, 95, 85 and 75% ethanol, each for 5 min. Samples were then stained in Harris hematoxylin for 4 min at room temperature and rinsed in tap water for 2 min. Differentiation was conducted using 1% hydrochloric acid, followed by washing with tap water. Slides were then counterstained with eosin for 2 min at room temperature. After eosin staining, the slices were dehydrated in 95 and 100% alcohol for 1 min at room temperature. The tissues were then stained with H&E and evaluated at x400 magnification under a light microscope (Olympus BX51).