Ab78494

Kidney tissues from mice were embedded in paraffin and 4 μm sections were used for Hematoxylin-eosin (HE) staining to detect morphological changes.
To perform immunohistochemical (IHC) staining, paraffin-embedded kidney tissue was deparaffinized in xylene, rehydrated, and blocked with 10% goat serum for 30 min. Rabbit polyclonal anti-KIM-1 (2.5 ug/ml, Abcam, ab78494) primary antibodies were incubated at 4°C overnight and subsequently incubated with HRP-conjugated secondary antibody for IHC.
Frozen sections of mouse’s kidney were cooled to room temperature for 10 min and then blocked with 5% goat serum (Beyotime, Shanghai, China) at room temperature for 1 h. Sections were washed with phosphate-buffered saline (PBS) followed by incubation with primary antibodies against DDIT3 (1:200 dilution, CST, 2895T) and SLC2A3 (1:100 dilution, Santa Cruz, sc74399) overnight at 4°C. After washing for three times with PBS, fluorescent secondary antibodies were added and incubated at room temperature for 1 h. After counterstaining with DAPI (Beyotime, Shanghai, China) for 10 min, sections were sealed under glass coverslips with an anti-fading fluorescence medium (Applygen, Beijing, China). Images were captured with a fluorescent microscope (Nikon, Tokyo, Japan).

Tissue sections were stained with hematoxylin and eosin (HE), Periodic acid-Schiff’s (PAS) and Masson’s trichrome stains using standard protocols. The mesangial area was determined from the PAS stained sections and the fibrotic area from the Masson’s trichrome stained sections as previously described^{28 (link)}. The antibodies used in this study were those against FXR (sc-13063, Santa Cruz), Grp78 (ab21685, Abcam), Chop (ab59396, Abcam), 4-hydroxynonenal (4-HNE, ab46545, Abcam), Kim-1 (ab78494, Abcam), 8-oxo-2′-deoxyguanosine (8-oxo-dG, ab64548, Abcam), SDHA (#11998, CellSignaling, Danvers, MA, USA), tumor necrosis factor (TNF, ab6671, Abcam), CD4 (sc-7219, Santa Cruz, Dallas, TX, USA), aSMA (NBP1-30894, Novus Biologicals, Littleton, CO, USA) and Collagen I (ColI, NB600-408, Novus Biologicals). TUNEL staining on kidney paraffin sections was performed with an ApopTag kit (Millipore, Billerica, MA, USA), according to the manufacturer’s instructions.