2.6. Immunohistochemical staining of BiP, CHOP and macrophages

Paraffin-embedded kidney sections were deparaffinized and incubated with 0.1 M sodium citrate, PH 6.0 at 65 °C for antigen retrieval. Then the slides were sequentially exposed to 3% H2O2 to block endogenous peroxidase activity, a buffer containing 5% BSA and 0.1% Triton X-100 to reduce non-specific binding, a specific primary antibody at 4 °C overnight, and HRP-conjugated secondary antibody for 1 h at room temperature. Signals of the antigen-antibody complexes were detected with a DAB kit following the protocol of the manufacturer. Finally, the slides were counterstained in hematoxylin. For quantification, 10–20 fields were randomly selected from each tissue section and the percentage of positive staining tubules or numbers of positive staining cells per mm² were evaluated.