Glioblastoma Cell Line Cultivation and Transfection

Human glioblastoma U251 MG (U251) and U87 MG (U87) cell lines were purchased from CLS Cell Lines Service (Germany) and monitored for correct genetic profile via microsatellite analyses according to a protocol described earlier (Peickert et al. 2012 (link)). LN-229 glioblastoma cells were from Prof. Marta Miączyńska from the International Institute of Molecular and Cell Biology in Warsaw, Poland. Cells were cultured at 37 °C and 5 % CO₂ in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with GlutaMAX-1, 10 % fetal bovine serum (FBS), 100 U/mL penicillin and 100 U/mL streptomycin (Gibco Live Technologies, USA). For most experiments, a formulated amino acid-free DMEM (Sigma-Aldrich, USA) was used. In order to prepare arginine- (-Arg) or lysine-free (-Lys) conditions, the medium was supplemented with all the amino acids except of arginine or lysine, respectively. A complete (control) medium was obtained by supplementation with all the amino acids (0.4 mM for arginine, and 0.8 mM for lysine and leucine). -Arg, -Lys and control media were supplemented with 5 % dialyzed FBS (Sigma-Aldrich, USA), lacking small molecules such as amino acids. U251 cells cultivated in the control medium were transfected with pmaxGFP plasmid using Nucleofection™ technology (Lonza Group Ltd., Switzerland).