Gentamycin

Cell culture materials were obtained from Greiner Bio-One International GmbH (Frickenhausen, Germany). If not otherwise indicated, cell culture solutions and supplements (L-glutamine, HEPES ((4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), penicillin, streptomycin, amphotericin B) and DPBS (Dulbecco´s Phosphate Buffered Saline) were obtained from Biochrom AG (Berlin, Germany). L-15 medium was purchased from Lonza Group AG (Visp, Switzerland) and Schneider´s Drosophila Medium (SDM) from Fisher Scientific GmbH (Schwerte, Germany). Sigma Aldrich (Taufkirchen, Germany) was used the supplier for chemicals such as galactose, lactalbumin hydrolysate, tetracycline, cell culture grade water (for medium preparation) and FCS (Fetal Calf Serum). Gentamycin was obtained from Biowest (Nuaillé, France). Ultrapure water for buffer preparation was produced by a Millipore unit (Synergy Water Purification System, Merck KGaA, Darmstadt, Germany). Collagenase type II from Clostridium histolyticum (CLS II, #C2-28, Lot Number 47N17872A, 280 U/mg) was purchased from Biochrom AG. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) reagent was received from Alfa Aesar (Ward Hill, Massachusetts, USA).
Sterile filters (0.2 µm cellulose acetate) were purchased from VWR International (Darmstadt, Germany) and cell strainers from pluriSelect Life Science (Leipzig, Germany).

AD293 cells were obtained from Agilent Technologies (United States). Cells were cultured in complete Dulbecco’s modified Eagle’s medium (Biowest, France) supplemented with 10% fetal bovine serum (FBS) (PAA, The Cell Culture Company, Austria), Penicillin-Streptomycin (Life Technologies, USA), Gentamycin (Biowest), Fungizone Antimycotic (Life Technologies), Primocin and Normocin (Invivogen, USA). Serum free conditions means a culturing in complete medium lacking FBS supplementation. Cultures were incubated at 37°C in humidified atmosphere and 5% CO₂ and passaged with Trypsin/EDTA (0.05%; Life Technologies). Cells were observed with Biostation CT (Nikon, Japan) and cell counting was performed with ImageJ (http://imagej.nih.gov/ij/) and CL-Quant Ver3.10 imaging software (Nikon).

DK-MG cells were purchased from Leibniz-Institut DSMZ (Braunschweig, Germany, Cat. no. ACC-277) and cultured in RPMI 1640 (GIBCO, Life Technologies, Grand Isle, NY, Cat. no. 11875-093). Obtaining of U87-MGvIII clone 4.12, DK-MG^high and DK-MG^low lines was described previously [28 (link)].
AD293 cells were purchased from Agilent Technologies (Santa Clara, CA, USA, Cat. no. 240085) and cultured in complete DMEM High Glucose (Biowest, France, Cat. no. L0102-500). Parental cell line was either transfected or transduced with lentiviral vectors carrying appropriate transgenes (prepared as described below) and positively selected using puromycin (5 μg/mL, InvivoGen, Cat. no. ant-pr-1).
The media used for culturing of cell lines were supplemented with 10% FBS (PAA, The Cell Culture Company, Austria, Cat. no. A15-104), 1% penicillin/streptomycin (Life Technologies, Carlsbad, CA, USA; Cat. no. 15140-122), 0.2% Gentamycin (Biowest, France, Cat. no. L0011-100). Cells were cultured under standard cell-culture conditions (5% CO₂, 16% O₂, 37°C).
Glioma cells were cultured as described previously [28 (link)]. Study was approved and conducted in accordance to the Approval No. RNN/234/17/KE from the Bioethical Committee of the Medical University of Lodz.