Paraffin-embedded liver Sects. (4 µm thick) were used for immunohistochemistry (IHC) experiments. The sections were dewaxed, rehydrated, and quenched with 3% H2O2, followed by heat-induced epitope retrieval in 10 mM citrate buffer (pH 6) at 95 °C for 20 min. Nonspecific antigens were blocked with 1% BSA (cat: A7906, Sigma-Aldrich). Anti-Ki67 (1:500, ab15580, Abcam) and anti- Hnf4α antibodies (1:500, 3113S, CST) were incubated overnight at 4 °C. Goat-anti-rabbit fluorescein isothiocyanate-labeled IgG or goat-anti-mouse rhodamine IgG (1:200, Proteintech, Rosemont, IL, USA) were incubated at 4 °C for 2 h, followed by 4′,6-diamidino-2-phenylindole (DAPI) staining (Abcam) the cell nucleus. Slides were mounted and visualized using an OLYMPUS microscope.
Primary hepatocytes or macrophages were seeded in a cell culture dish for immunofluorescence experiments. The cells were fixed and permeabilized at 4 °C for 30 min. After incubation with anti-Tet2 (1:100, ab124297, Abcam), anti-Ki67 (1:100, ab15580, Abcam), and anti-Stat1 antibodies (1:100, 9176, CST) at 4 °C overnight, the cells were washed with PBS and stained with goat-anti-rabbit FITC-labeled IgG or goat-anti-mouse rhodamine IgG (1:200, Proteintech) at 4 °C for 2 h, followed by DAPI staining (Abcam). The cells were viewed using a Zeiss Confocal Microscope Imaging System (Carl Zeiss, Jena).
Primary hepatocytes or macrophages were seeded in a cell culture dish for immunofluorescence experiments. The cells were fixed and permeabilized at 4 °C for 30 min. After incubation with anti-Tet2 (1:100, ab124297, Abcam), anti-Ki67 (1:100, ab15580, Abcam), and anti-Stat1 antibodies (1:100, 9176, CST) at 4 °C overnight, the cells were washed with PBS and stained with goat-anti-rabbit FITC-labeled IgG or goat-anti-mouse rhodamine IgG (1:200, Proteintech) at 4 °C for 2 h, followed by DAPI staining (Abcam). The cells were viewed using a Zeiss Confocal Microscope Imaging System (Carl Zeiss, Jena).