Anti bcl3

IHC analysis was performed using the following antibodies: rabbit anti-p21 (diluted 1:100, Abcam, Cambridge, UK), anti-Bcl₃ (diluted 1:100, Abcam, Cambridge, UK), anti-Bcl3 (diluted 1:100, Protechtein) anti-IL8 (diluted 1:100, Abcam, Cambridge, UK), anti-CD68 (diluted 1:100, Abcam, Cambridge, UK), anti-iNOS (diluted 1:100, Abcam, Cambridge, UK), and anti-CD163 (diluted 1:200, Abcam, Cambridge, UK). The detailed method has been published previously [42 (link)]. Five fields in each section were randomly selected to calculate the ratio of positive expression area. For the analysis, the positive intensity of IHC is calculated by counting the number of positive cells in the field of random collection for each marker. For IF, rabbit anti-HNF-4α (diluted 1:50, Abcam, Cambridge, UK), anti-p21 (diluted 1:50, Abcam, Cambridge, UK), anti-Bcl3 (diluted 1:50, Abcam, Cambridge, UK), and anti-IL8 (diluted 1:50, Abcam, Cambridge, UK) were used.

Tissues or cultured cells were lysed with RIPA lysis buffer with protease inhibitors and phosphatase inhibitors (Beyotime, Shanghai, China) using standard methods. After centrifugation and quantification, 25 µg of protein was loaded for blotting with the indicated specific antibodies. A NE-PER Nuclear and Cytoplasmic Extraction kit (Thermo Fisher Scientific, Waltham, USA) was used to isolate nuclear and cytoplasmic fractions according to the manufacturer’s protocol. The following antibodies were used: anti-Sox9 (1:1000, Abcam, Cambridge, UK), anti-HNF4α (1:2000, Abcam, Cambridge, UK), anti-TERT (1:1000, Novus, Colorado, USA), anti-E-Cadherin (1:1000, Abcam, Cambridge, UK), anti-Vimentin (1:1000, CST, Danvers, USA), anti-CK19 (1:1000, Abcam, Cambridge, UK), anti-LGR5 (1:1000, Abcam, Cambridge, UK), anti-Epcam (1:1000, Abcam, Cambridge, UK), anti-P65 (1:1000, CST, Danvers, USA), anti-p-P65 (1:1000, CST, Danvers, USA), anti-Bcl3 (1:100, Abcam, Cambridge, UK), anti-Histone H3 (1:1000, Proteintech, Rosemont, USA), anti-YAP1 (1:1000, CST, Danvers, USA), anti-CTGF (1:1000, Abcam, Cambridge, UK), anti-Cyr61 (1:1000, Abcam, Cambridge, UK), anti-His (1:5000, Proteintech, Rosemont, USA), anti-Flag (1:2000, Proteintech, Rosemont, USA), anti-Myc (1:1000, Proteintech, Rosemont, USA), anti-Ubiquitin (1:1000, CST, Danvers, USA), and anti-β-actin (1:4000, Bioworld, MN, USA).

Whole-cell proteins were extracted using RIPA lysis buffer supplemented with protease inhibitors (50 mM Tris–HCl, pH 7.4, 1% Nonidet P-40, 0.25% SDS, 150 mM NaCl, 1 mM EDTA, 1mM PMSF, 1mM NaF, 1mM Na₃VO₄, 2 µg/ml aprotinin, 1 µg/ml pepstatin, and 1 µg/ml leupeptin). Lysates were resolved on SDS-PAGE gels, transferred to nitrocellulose membranes and immunoblotted with anti-p105/p50 (Cell Signaling Technology), anti-BCL-3 (AbCam).