Nch421k

Cell line U-87 MG was obtained from American Type Culture Collection (ATCC) and was cultured in high glucose DMEM supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine and 1% penicillin/streptomycin. Glioblastoma stem cell line NCH421k was purchased from Cell Lines Service (CLS) GmbH and cultured in Neurobasal Medium (Invitrogen, Waltham, MA, USA) supplemented with 2 mM L-glutamine, 1 × B-27 supplement (Invitrogen), 1 U/mL heparin (Sigma-Aldrich, St. Louis, MO, USA), 20 ng/mL bFGF and EGF (both from Invitrogen, Life Technologies, Carlsbad, CA, USA) and 1% penicillin/streptomycin. For in vivo assay, cells were transfected with plasmid vectors pCMVDsRed-Express2 and pEGFP-N1 to stably express red fluorescent protein DsRed and green fluorescent protein EGFP, as described previously [31 (link),32 (link)]. All cell lines were cultured at 37 °C, 5% CO₂ and 95% humidity and were tested for mycoplasma contamination using the MycoAlert Mycoplasma Detection Kit (Lonza, Basel, Switzerland).

Human glioblastoma cell lines U373 (cell line listed under catalogue number 89,081,403 has been re-named as U-251) and U87 were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA) and were cultured in DMEM high glucose medium (GE Healthcare, Chicago, IL, USA), supplemented with 10% (v/v) FBS, 2 mM l-glutamine, 100 IU/mL penicillin and 100 µg streptomycin. Glioblastoma stem cell line NCH421k was obtained from CLS (Cell Lines Service GmbH, Eppelheim, Germany) and expanded as spheroid suspensions in complete Neurobasal Medium (Invitrogen, Life Technologies, Carlsbad, CA, USA) containing 2 mM l-glutamine, 1 × penicillin/streptomycin, 1 × B-27 supplement (Invitrogen, Life Technologies, Carlsbad, CA, USA), 1 U/mL heparin (Sigma-Aldrich, St. Louis, MO, USA), 20 ng/mL bFGF and EGF (both from Invitrogen, Life Technologies, Carlsbad, CA, USA). Cells were transfected with the plasmid vector pCMVDsRed-Express2 and pEGFP-N1 to stably express the red fluorescent protein DsRed and enhanced green fluorescent protein eGFP, as described previously [59 (link),63 (link)]. All cell lines were maintained at 37 °C with 5% CO₂ and 95% humidity. All cell cultures were tested for mycoplasma contamination using the MycoAlert Mycoplasma Detection Kit (Lonza, Basel, Switzerland).