Rpmi 1640 growth media

Bovine serum albumin (BSA), biotinylated bovine serum albumin (bio-BSA), streptavidin, streptavidin-Cy3 conjugate (SA-Cy3), triethanolamine (TEA), Zeba 7K Da molecular weight cutoff desalting spin columns, and Fluosphere fluorescent nanoparticles (20 nanometer diameter, carboxylate-terminated, nile red fluorescence) were purchased from Thermo Fisher Scientific. Phosphate buffered saline (PBS), fluorescein isothiocyanate isomer I, eosin-5-isothiocyanate (EITC), Cy3 NHS ester, poly(ethylene glycol) diacrylate (M_n~575 Da), monomethyl ether hydroquinone dehibiting columns, vinyl-2-pyrrolidinone, thiazoyl blue tetrazolium bromide (MTT), and dimethyl sulfoxide (DMSO) were purchased from Sigma Aldrich. Epoxy-functionalized, glass microarray slides were supplied by Array-It Corp. A non-small cell lunger cancer line A549 was purchased from ATCC. RPMI-1640 growth media, penicillin/streptomycin, and trypsin/EDTA (0.25% solution) were purchased from Gibco. Advantage grade fetal bovine serum (FBS) was purchased from Atlanta Biologicals. The primary antibody, anti-human EGFR was purchased from Biolegend. The secondary antibody, FITC-labeled horse anti-mouse IgG was purchased from Vector Labs.

Parental Ba/F3 cells were obtained from DSMZ (Braunschweig, Germany). Ba/F3 cells were cultured under standard conditions in RPMI 1640 growth media (Gibco, Karlsruhe, Germany) containing 10% fetal bovine serum, 200 U penicilin per mL, and 200 g streptomycin per mL (Gibco, Karlsruhe, Germany). IL3-dependend Ba/F3 cells were supplemented with 2 ng/mL interleukin-3 (IL-3; R&D, Wiesbaden, Germany). Ba/F3 p185wt cells were infected with retrovirus (pBabe-puro, pBabe-AurA, pBabe-AurB) and puromycin-treated for selection. For synchronisation in G_0/1, cells were serum starved for 72 h in the presence of 0,5% FCS.

The K1 cell line was obtained from health protection agency culture collection, BCPAP cell line was purchased from Chinese Academy of Science. BHP 2-7 cell line was kindly provided by Prof. Jerome M. Hershman. All cell lines were maintained at 37°C and 5% CO₂ in humidified atmosphere and grown in RPMI 1640 growth media supplemented with 10% fetal bovine serum (GIBCO). Under these culture conditions, cells were treated with dabrafenib, selumetinib and lapatinib (MCE) individually or in combination for indicated times. Dimetylsulfoxide (DMSO) was used in parallel as vehicle control.

Human clear-cell RCC cell lines RCC-FG1/KTCTL-26, RCC-MF/KTCTL-1M (both from Cell Line Service (CLS), Eppelheim, Germany) and 769-P (from ATCC; American Type Culture Collection USA) were cultured in RPMI1640 growth media (Gibco, ThermoFischer Scientific, Vienna, Austria) containing 10% fetal bovine serum (Hyclone, GE Healthcare, Frankfurt/Main, Germany) and 1% penicillin/streptomycin mixture (Gibco). Human umbilical vein endothelial cells (HUVECs) were cultured in EBM™-2-Medium with necessary supplements (EGM™-2 SingleQuots™; all purchased from Lonza, Basel, Switzerland). All cell lines were kept at 37 °C in a humidified 5% CO₂ atmosphere.

The HT-115 (human colon carcinoma cells) was obtained from American Type Culture Collections (ATCC), Manassas, Virginia, USA. Vero (kidney epithelial cells) and V79 (hamster lung fibroblast cell) cell lines were purchased from the National Center for Cell Sciences (NCCS), Pune, India. RPMI-1640 growth media, fetal bovine serum (FBS), penicillin and penicillin/streptomycin, and cell-culture-associated chemicals were purchased from Gibco, Paisley, UK. The cDNA synthesis kit was purchased from Qiagen, Hilden, Germany. SYBR Green PCR Master Mix was purchased from Qiagen, CA, USA. Propidium iodide, acridine orange, ethidium bromide, dimethyl sulfoxide, and trypsin were obtained from Sigma-Aldrich chemical company (St. Louis, MO, USA).

The hepatoblastoma cell lines HuH-6 (RRID:CVCL_4381), HepT1 (RRID:CVCL_G003), Hep-T3 (RRID:CVCL_G004), and HepG2 (RRID:CVCL_0027) were cultured in RPMI 1640 growth media (Gibco, Thermo Fischer Scientific, Germany), supplemented with 10% fetal calf serum, 100 U/mL penicillin, and 100 μg/mL streptomycin, at 37 °C in a humidified chamber with a saturated atmosphere containing 5% CO₂. Cells were passaged at a confluency of 80–90% with 0.05% trypsin (v/v) and 0.2% EDTA (w/v) (Sigma-Aldrich, Germany) in Dulbecco’s phosphate-buffered saline (PBS). For gene expression and methylation analysis Huh-6 and Hep-T3 cells were treated with 0.5 µM 5-aza-2′-deoxycytidine (5-aza; Sigma-Aldrich, Germany), HepT1 and HepG2 cells with 1.25 µM 5-aza or solvent for 3 and 5 days.