Pfk118 310

PFK118–310 was purchased from Sigma-Aldrich while ICG-001 was obtained from Selleck Chemicals. Alexa fluor 488 phalloidine and 4′, 6-diamidino-2-phenylindole (DAPI) were purchased from Sigma. The antibodies used for Western blot analysis and immunocytochemistry were: rabbit anti-CD44 (Abcam # 51037), rabbit anti-CD133 (Cell Signaling # 3663), mouse anti-CD166 (Abcam # 175422), rabbit anti-Lgr5 (Sigma-Aldrich # HPA012530), goat anti-actin (Santa Cruz # sc-1615), rabbit anti-beta-catenin (Cell Signaling # 8480), mouse antibodies specific for the active form of beta-catenin, dephosphorylated on Ser 37 and Thr 41 (Millipore, clone 8E7, # 05–665). Anti-rabbit IgG horseradish peroxidase-linked antibodies and anti-mouse IgG horseradish peroxidase-linked antibodies were from Cell Signaling whereas Cy3-conjugated anti-mouse secondary antibodies were obtained from Jackson ImmunoResearch Labs.
The following antibodies were used for flow cytometry analysis: phycoerythricine-conjugated mouse anti-human ABCB1 (BD Biosciences # 557003), phycoerythricine-conjugated mouse anti-human ABCG2 (BD Biosciences # 561180), mouse anti-human ABCC1 (BD Biosciences # 557594) and anti-mouse phycoerythricine-conjugated secondary antibodies (BD Biosciences # 555574).

The human hepatocellular carcinoma (HepG2 and Hep3B) and human embryonic kidney (HEK293T) cell lines were kindly provided by Professor Claude Szpirer, Université Libre de Bruxelles, Belgium. The human colon adenocarcinoma (SW480) cell line was purchased from CLS (Germany). HepG2, Hep3B, SW480 and HEK293T cells were cultured in advanced DMEM medium (Invitrogen) supplemented with 10% fetal bovine serum (Biowest), 2 mM L-glutamine, 50 units/ml penicillin, and 50 μg/ml streptomycin. Cells were maintained in an incubator with humidified air (5% CO2) at 37°C. PFK118-310 (#K4394) was purchased from Sigma and used at a 0,2 or 0,4μM concentration from a DMSO solution.