Immunohistochemical Analysis of Kidney Samples

After dewaxing and rehydration, slides were incubated with 3% H₂O₂ for quenching endogenous peroxidase activity and then heated in boiling antigen unmasking solution for antigen retrieval. Slides were blocked with 5% goat serum (Vector Laboratories, UK) for 1 h, then incubated with primary antibody in a humidified chamber at 4℃ overnight. Positive signal was visualized by DAB solution (ZSGB-BIO, Beijing, China). The images were visualized and acquired using the Leica DM 1000 microscope image system. Quantitative evaluation involved the use of Image-Pro Plus 6.0 (Media Cybernetics, Rockville, MD, USA) 58 (link). Positive signals and the number of positive cells in the kidney were quantified in 10 bright fields (×200) in the eyepiece of the microscope.