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Kidney tissues excised from each mouse were fixed in 10% buffered formaldehyde immediately after removal from the animals. Fixed tissues were routinely processed, then embedded in paraffin, and cut into 4 μm thick sections; they were mounted on slides for hematoxylin and eosin staining. Qualitative examinations of prepared tissues and the obtaining of their photos were carried out using a light microscope (Olympus BX61- USA) connected to motorized controller unit (Olympus bx-ucb, USA) and photographed by a camera (Olympus DP72, USA).

At the end of 4 weeks animals were euthanized under deep ether anaesthesia. Abdomen and pelvic cavities were opened, to remove the testis. It was immersed in 10 % neutral buffered formalin 24 h after which it cut transversely and processed for paraffin sections 5-μm thick stained by hematoxylin and Eosin (H&E) for general structure examination using a light microscope (Olympus BX61- USA) connected to motorized controller unit (Olympus bx-ucb, USA) and photographed by a camera (Olympus DP72, USA).
For Ki 67 immunostaining, “another serial sections of 5-μm paraffin tissue were cut, dewaxed, and rehydrated in xylene and distilled water. Antigen retrieval was then carried out in a microwave oven. 3 % hydrogen peroxide was used to quench endogenous peroxidase activity in sections for 30 min. at room temperature, followed by 15 min. of blocking with 5 % bovine serum. The sections were then incubated overnight at 4 °C in a humidified room with specific primary antibodies against Ki-67 (GB11010; Wuhan Saiweier Biological Technology, China). After being washed with PB, the sections were treated with secondary antibody and stained with 3, 3′-diaminobenzidine after being rinsed with PB. Hematoxylin was used to counterstain the slices, which were then washed in tap water”. Micrographs of (IHC) were examined under a microscope (Zhao et al, 2018 (link)).