Anti α tubulin clone 1e4c11

Mouse monoclonal anti-ABCB4 (clone P3-II-26) and anti-MRP2 (multidrug resistance-associated protein 2; clone M2-I-4) antibodies were purchased from Enzo Life Sciences (Villeurbanne, France); and anti-α-tubulin (clone 1E4C11) was sourced from ProteinTech (Manchester, United Kingdom). Alexa Fluor-labeled secondary antibodies, the DRAQ5 fluorescent probe and culture media came from ThermoFisher (Cergy-Pontoise, France), and peroxidase-conjugated secondary antibodies were acquired from Rockland Immunochemicals (Gilbertsville, PA, USA). The ECL-Prime detection kit was from VWR (Courtaboeuf, France). The transfection reagents Turbofect and JetPrime were purchased from Thermo-Fisher Scientific, (Saint-Herblain, France) and Ozyme (Saint-Cyr-l’Ecole, France), respectively. Ivacaftor (VX-770) came from Clinisciences (Nanterre, France). All other reagents were obtained from Sigma-Aldrich (Lyon, France).

GFP-tagged Abcb11-encoding vectors (wt, A257V, T463I or G562D) were transiently transfected in human hepatocellular carcinoma HepG2 cells as published [34 (link)]. These cells form pseudo-bile canaliculi in culture and allow localization studies [35 (link)]. MDCK cells were stably transfected as described [10 (link)]. MDCK clones with the highest Abcb11-GFP (wt, A257V, T463I or G562D) expression and parental MDCK cells were infected with Ntcp- encoding lentiviral particles as described [10 (link)]. Immunofluorescence and immunoblotting analyses were performed as described [10 (link)] using the following primary antibodies: rat monoclonal anti-cMyc (Clone JAC6; GeneTex, Irvine, CA, USA), rabbit polyclonal anti-GFP (ab290; Abcam, Cambridge, UK), mouse monoclonal anti-ABCC2 (clone M2I-4; Enzo Life Sciences, Villeurbanne, France), mouse monoclonal anti-GFP (clone 7.1 and 13.1; Roche Diagnostics, Mannheim, DE, USA), anti-cMyc (Clone 9E10; BD Biosciences Pharmingen, San Diego, CA, USA) and anti-α-tubulin (clone 1E4C11; ProteinTech, Manchester, UK). Peroxydase- and fluorochrome-conjugated secondary antibodies were from GE Healthcare (Chicago, IL, USA) and Molecular Probes/Thermo Fisher Scientific (Illkirch, France), respectively.