Fbs rpmi 1640

293T, A549, H460, and H1299 cells were obtained from the American Type Culture Collection (USA) (Authenticated Nov, 2018 by STR sequencing). p53^+/+- and p53^−/−-HCT116 cells were kindly provided by Dr. Yu-Ju Chen (Institute of Chemistry, Academia Sinica, Taiwan). CL1-5 cells were kindly provided by Dr. Pan-Chyr Yang (Graduate Institute of Oncology, National Taiwan University Medical College, Taiwan). 293 T, A549, H460, p53^+/+- and p53^−/−-HCT116 cells were cultured in 10% FBS/DMEM (Gibco, NY, USA); H1299 and CL1-5 cells were cultured in 10% FBS/RPMI-1640 (Gibco). Cells were passaged every 3 days in a 0.5% trypsin-EDTA solution and maintained until used.

LLC and A549 cells were purchased from ATCC (Manassas, VA, USA), and A549^EGFP cells were generated in our laboratory from A549 cells as previously described^{9 (link)}. These cells were maintained in 10% FBS RPMI 1640 (Gibco, Waltham, MA, USA). hLECs were purchased from ScienceCell and cultured in 10% FBS endothelial cell medium (ECM, ScienceCell Research Laboratories, Carlsbad, CA, USA).

The Mab 4D1 and Mab 8D6 hybridoma clones were cultured in 5% (v/v) FBS-RPMI 1640 (Gibco). The Mab 4D1 (murine IgG) binds a novel T. marneffei yeast phase-specific mannoprotein antigen and the Mab (29 , 30 (link)) was purified by HiTrap column protein G affinity chromatography (GE Healthcare) as described (30 (link)). In contrast, the HiTrap IgM affinity column (2-mercaptopyridine coupled to cross linking epharose) was chosen to purify Mab 8D6 (murine IgM), which is a Mab specific to melanin (31 (link)). All experiments were carried out as per the manufacturer’s instructions.

HUVECs were cultured in EGM (endothelial cell growth media)-2 BulletKit supplemented with SingleQuots (endothelial basal media-2 (EBM-2) supplemented with growth factors, cytokines, antibiotics; Lonza)/10% (v/v) FBS). HMVECs were cultured in EGM-MV BulletKit supplemented with SingleQuots (EBM-2 supplemented with growth factors, cytokines, antibiotics; Lonza)/20% (v/v) FBS. HT-29 (colon cancer cells) and MCF-7 cells (breast cancer cells) were cultured in 10% (v/v) FBS/DMEM with 1% (v/v) penicillin–streptomycin (5000 U/mL; Life Technologies). For PC3 cells (prostate cancer cells), 10% (v/v) FBS/RPMI 1640 media with 1% (v/v) penicillin–streptomycin (5000 U/mL; Life Technologies) was used during culture. All cells were maintained at 37°C in 5% CO₂.

The cell lines used in this study were purchased from ATCC, USA. Lung cancer cell lines A549, H23, H661 and H838 were grown in 10% FBS-RPMI 1640 (Life Technologies, CA) in 5% CO₂ cell culture incubator. The reagents L-glutamine and L-diazo-5-oxo-L-norleucine (DON) are purchased from Sigma-Aldrich, USA. As described elsewhere [60 (link), 66 (link), 67 (link)], for glutamine studies, the cells were plated in 6-well tissue culture plates at a density of 0.2 million cells/well. The following day, the cells were washed twice with serum free glutamine-free medium and replaced with 10% dialyzed FBS containing glutamine-free RPMI 1640 (Life Technologies, CA), and then incubated for 48 h. The corresponding glutamine-replete medium was prepared by addition of indicated concentration of glutamine. For DON (a glutamine analogue) studies, we added indicated concentration of DON to the cells which are already in 10% dialyzed FBS + 2 mM Gln+ RPMI 1640 medium [68 (link), 69 (link)]. Bronchial lung epithelial cells, BEAS-2B were grown in BEBM medium with supplements (Lonza, USA). Adenoviruses and lentiviruses were generated by the University of Michigan Vector Core. Lung cancer cells were infected with lentiviruses expressing PPAT shRNAs, PAICS shRNAs or Non-target controls only, and stable cell lines were generated by selection with 1 μg/ml puromycin (Life Technologies, CA).

Epithelial ovarian cancer (EOC) tissue samples and normal ovarian tissues were acquired from surgical specimens of patients between May 2019 and June 2022 at Wuhan University Zhongnan Hospital. This research study was authorized by the Wuhan University Zhongnan Hospital Ethics Committee (KELUN2021087) according to the principles outlined in the Declaration of Helsinki. The hospital pathology service validated the pathologies in the EOC tissue samples. Shanghai Zeye Bioengineering (Shanghai, China), and Procell Life Science & Technology (Wuhan, China), were all used to obtain OC cell lines ES-2, Hey, OVCAR-3, SKOV3, A2780, and the human SV40-transformed immortal cell line IOSE80. ES-2, Hey, OVCAR-3, SKOV3, and A2780 cells were cultured in DMEM (Life Technologies, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (FBS; BI, Kibbutz, Israel). IOSE80 cells were grown in 10% FBS RPMI1640 (Life Technologies). These cells were cultured in a 37 °C incubator with 5% CO₂. Cell lines were validated by STR profiling, and there was no mycoplasma contamination.