Ab220788

Total protein was separated from the cell lysis buffer. For the detection of β-catenin in nuclei and cytoplasm, cells were collected and used to extract nuclear proteins and cytoplasmic protein, respectively (Biotechnology, Shanghai, China).The protein concentration was measured using the bicinchoninic acid (BCA) method (Beyotime). Proteins (20 µg) were separated via SDS polyacrylamide gels and then transferred onto PVDF membranes. After blocking for 2 h with 5% nonfat milk at room temperature, these membranes were incubated overnight with primary antibodies (MUC5AC, ab198294, 1/20,000; ICAM-1, ab282575, 1:1000; Bcl-2, ab32124, 1:1000; Bax, ab32503, 1:1000; cleaved caspase3, ab32042, 1:500; cleaved caspase9, ab2324, 1:1000; MMP10, ab261733, 1:1000; GAPDH, ab8245, 1:5000; β-catenin, ab68183, 1:1000; β-actin, ab179467, 1:5000; TBP, ab220788, 1:1000; Abcam, England). Membranes were washed three times by PBS and then incubated for 2 h at room temperature. The protein bands were visualized using an enhanced chemiluminescence (ECL) kit (Thermo Fisher Scientific, Inc.).

Custom-made mouse monoclonal CTCF antibodies were previously described. [96 (link)]. Custom-made mouse monoclonal BORIS antibodies were also described previously [22 (link)]. Rabbit polyclonal antibody against SRCAP (Kerafast, ESL103), rabbit polyclonal against HCFC1 (Novus Biologicals, NB100-68209), goat affinity-purified polyclonal antibody against Mxi1 (R&D Systems, AF4185), rabbit monoclonal to TATA-binding protein TBP (Abcam, ab220788), rabbit polyclonal anti-PHF8 antibody (Bethyl Laboratories, A301-772A), rabbit polyclonal to MAZ (Abcam, ab85725), rabbit polyclonal antibody against the region of histone H3 containing the trimethylated lysine 27 (H3K27me3) (Diagenode, C15410195), rabbit polyclonal antibody against histone H3, trimethylated at lysine 36 (H3K36me3) (Diagenode, C15410058), rabbit polyclonal antibody against histone H2A.Z (Abcam, ab4174), rabbit polyclonal to Histone H3 (acetyl K27) (Abcam, ab4729), rabbit polyclonal to Histone H3 (tri methyl K4) (Abcam, ab8580), anti-RNA Polymerase II Antibody, CTD Antibody, clone 8WG16 (Millipore, 05–952-I-100UG).