Mem non essential amino acid solution

The estrogen receptor (ER)-positive human breast cancer line MCF-7 (RCB1904) and the ER-negative human breast cancer cell line MDA-MB-453 (RCB1192) were purchased from the RIKEN cell bank. MCF-7 cells were cultured at 37 °C with 5% CO₂ in E-MEM with L-glutamine and phenol red (Wako Pure Chemical Industries, Ltd., Osaka, Japan) containing 10% fetal bovine serum (GE Healthcare Life Sciences, Inc., Logan, UT, USA), 1% penicillin/streptomycin (Wako Pure Chemical Industries, Ltd.), 1.0 mM sodium pyruvate solution (Wako Pure Chemical Industries, Ltd.), and 1% MEM non-essential amino acid solution (Thermo Fisher Scientific, Inc., Waltham, MA, USA). MDA-MB-453 cells were cultured at 37 °C without CO₂ in Leibovitz′s L-15 Medium with L-glutamine, phenol red, and sodium pyruvate (Wako Pure Chemical Industries, Ltd.) containing 10% fetal bovine serum (GE Healthcare Life Sciences, Inc.) and 1% penicillin/streptomycin (Wako Pure Chemical Industries, Ltd.).
The hyperthermia treatment was based on our previous study [11 (link)]. The MCF-7 cells were incubated at 42 °C with 5% CO₂ for 1 h using a standard incubator, and the MDA-MB-453 cells were incubated in the same way, but without CO₂.

Vero cells and KMB17 cells obtained from the Institute of Medical Biology, Chinese Academy of Medical Sciences, and Peking Union Medical College, China, were maintained in minimum essential medium (MEM) (HyClone) supplemented with 10% fetal bovine serum (FBS) (Gibco) and 1% penicillin/streptomycin at 37°C with 5% CO₂. 293 T cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with 10% FBS and 1% penicillin/streptomycin at 37°C with 5% CO₂. T98G cells were cultured in modified MEM supplemented with 10% FBS, 1 mM sodium pyruvate, 1×MEM nonessential amino acid solution (Thermo Fisher Scientific) and 1% penicillin/streptomycin at 37°C with 5% CO₂. Human EV-A71 strain 87-2008 Xi’an Shaanxi (GenBank ID: HM003207.1) was kindly supplied by Professor Wenbo Xu in Chinese Centre for Disease Control and Prevention. The EV-A71 infection was performed in Vero cells, KMB17 cells and T98G cells. The titers of EV-A71 and its mutant derivatives were determined by plaque assay in Vero monolayers.

Human lymphoblastoid HLA Class I-reduced cell lines (C1R) overexpressing HLA-B*57:01 or HLA-B*15:02, were cultured in Roswell Park Memorial Institute (RPMI) media (Thermo Fisher Scientific, MA, USA) supplemented with 2 mM Minimum Essential Medium (MEM) non-essential amino acid solution (Thermo Fisher), 100 mM (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) (ICN Biochemicals, Aurora, OH, USA), 2 mM l-glutamine (Merck, Darmstadt, Germany), 0.6 mg/L benzylpenicillin (CSL, Melbourne, Australia), 1 mg/L streptomycin (CSL), 50 µM 2-mercaptoethanol (Merck) and 10% FBS (CSL, Melbourne) with 1% G418 selection antibiotic (Thermo Fisher) and were kept in humid conditions at 37 °C, 5% CO₂.

Primary human foetal cortical normal astrocytes were a gift from Prof. DK Male (The Open University, Milton, Keynes, UK) and were cultured as described elsewhere [47 (link)]. The VUMC-DIPG-A (H3.3 K27M) cell line was kindly provided by Dr. Esther Hulleman (VUMC Cancer Center, Amsterdam, the Netherlands) [22 (link)]. SU-DIPG-IV (H3.1 K27M) cells were kindly provided by Dr. Michelle Monje (Stanford University, California, USA). [48 (link)]. VUMC-DIPG-A cells were cultured in 1:1 DMEM-F12 and Neurobasal-A cell media and supplemented with a 1% (v/v) glutamax supplement, a 1% (v/v) antibiotic–antimycotic solution, 10 mM HEPES, a 1% (v/v) MEM non-essential amino acid solution, and 1 mM sodium pyruvate (Thermofisher, Paisley, UK), hereafter referred to as TBM medium. VUMC-DIPG-A medium was also supplemented with 10% heat-inactivated foetal bovine serum (Merk, Watford, UK). SU-DIPG-IV was cultured in TBM medium supplemented with a 2% (v/v) B27 supplement without vitamin A (Thermofisher), 20 ng/mL of bGFG, 20 ng/mL of EGF, 10 ng/mL of PDGF-AA, 10 ng/mL of PDGF-BB (Peprotech), and 5 IE/mL of heparin (Merk). Cells were grown in adherence and passaged using 0.25% (v/v) trypsin-EDTA (Merk). All cells were cultured at 37 ℃ in a humidified environment containing 5% CO₂.

The cell line, 293GP, from Riken Cell Bank, was maintained in Dulbecco’s Modified Eagle Medium (DMEM; Wako), supplemented with 10% fetal bovine serum (FBS) and MEM non-essential amino acid solution (Thermo Fisher Scientific). 293T (ATCC), RAW264.7 (ATCC), and Huh7.5.1 (a gift from Dr. Francis V. Chisari) cells were maintained in DMEM supplemented with 10% FBS.