Anti tubulin 2148

The following antibodies and reagents were purchased for use in the study: anti-α-smooth muscle actin antibody (α-SMA; A5228) from Sigma-Aldrich (St Louis, MO, USA); anti-MMP2 (#40994), anti-GAPDH (#5174), and anti-Tubulin (#2148) antibodies and normal rabbit IgG (#2729) from Cell Signaling Technology (Danvers, MA, USA); anti-COL1A1 (PAB17205), anti-Fibronectin (ab2413), anti-Timp1 (ab211926), and anti-TGFβ1 (ab179695) antibodies from Abnova (Colorado, USA) and Abcam; and TGFβ1 (240-B) from R&D Systems (Minneapolis, MN, USA). In addition, ABI Taqman primers/probes were purchased from Applied Biosystems (Foster City, CA, USA).

Western blot analysis was performed as previously described^{53 (link)}. The following antibodies were used: anti-HDAC6 (sc-11420, Santa Cruz), anti-HDAC10 (H3413, Sigma), anti-acetylated histone H3 (06-911, Merck Millipore), anti-histone H3 (9715, Cell Signaling Technology, Cambridge UK), anti-LAMP-1 (H4A3, DHSB, Iowa City, IA, USA), anti-LAMP-2 (H4B4; Santa Cruz Biotechnology), anti-acetylated tubulin (6–11B-1; Sigma), anti-tubulin (2148, Cell Signaling Technology), anti P-glycoprotein (13978 S, Cell Signaling Technology) and anti-β-actin (clone AC-15; Sigma).

Precipitated proteins were eluted with for immunoblotting, and cells were lysed in RIPA buffer (50 mM Tris-HCl pH 7.4; 1% NP-40; 0.5% Na deoxycholate; 0.1% SDS; 150 mM NaCl; 2 mM EDTA; PIC and PhIC) and centrifuged at 12,000 rpm, 4 °C. The protein concentration was measured using a Qubit protein assay kit (Thermo Fisher Scientific, Waltham, MA, USA) and mixed with 4× Laemmli buffer (200 mM Tris–HCl pH 6.8; 4% SDS; 40% glycerol; 0.01% bromophenol blue; 100 mM DTT) and then boiled for 10 min. After electrophoresis, the proteins were transferred to the nitrocellulose membrane (Bio-Rad, Hercules, CA, USA) and stained with 5% milk (Cell Signaling Technology, Danvers, MA, USA) in PBS. Endogenous PHF10 isoforms were detected by anti-PHF10 antibodies described previously [28 (link)]. These polyclonal antibodies were generated in our laboratory by immunizing rabbits with the His-tagged polypeptide corresponding 89–370 amino acids (NP_060758.2) contained in all four PHF10 isoforms. The M2 clone (Sigma-Aldrich, Saint Louis, MO, USA) was used as anti-FLAG antibodies. Anti-Tubulin (#2148) and anti-GAPDH (14G10) antibodies were from Cell Signaling. HRP-conjugated anti-rabbit IgG and HRP-conjugated anti-mouse goat IgG were from DHGB.