Procedures have recently been described [2 (link)]. The patented KGN cell line was obtained from RIKEN BioResource Center [4 (link)] with permission by T. Yanase. KGN cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM)/Ham’s F12 medium (Life Technologies, Paisley, UK) supplemented with penicillin (100 U/mL), streptomycin (100 μg/mL) (Biochrom, Berlin, Germany) and 10% fetal calf serum (FCS) (Capricorn Scientific, Ebsdorfergrund, Germany) at 37 °C and with 5% CO2. For stimulation experiments, 20 µM Trolox (Santa Cruz Biotechnology, Dallas, TX, USA) or 100 µM/1 mM H2O2 (Sigma-Aldrich, St. Louis, MO, USA) was diluted in DMEM/Ham’s F12 medium (Life Technologies; colorless medium without phenol red was used for live cell fluorescence imaging to reduce background autofluorescence).
Dmem ham s f12 medium
Manufactured by Thermo Fisher Scientific
Sourced in United States, United Kingdom, Germany, France
DMEM/Ham's F12 medium is a basal medium commonly used for the cultivation of a variety of cell types. It is a combination of Dulbecco's Modified Eagle's Medium (DMEM) and Ham's F12 nutrient mixture, providing a balanced formulation of amino acids, vitamins, and other essential components to support cell growth and maintenance in cell culture applications.
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Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (26 mentions)
Streptomycin, by Thermo Fisher Scientific (16 mentions)
Penicillin, by Thermo Fisher Scientific (12 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (12 mentions)
Non essential amino acids (neaa), by Thermo Fisher Scientific (7 mentions)
L glutamine, by Thermo Fisher Scientific (7 mentions)
Mcf 7, by Thermo Fisher Scientific (6 mentions)
Trypsin edta, by Thermo Fisher Scientific (6 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (5 mentions)
Insulin, by Thermo Fisher Scientific (5 mentions)
Insulin, by Merck Group (5 mentions)
Hydrocortisone, by Merck Group (4 mentions)
2 mercaptoethanol, by Thermo Fisher Scientific (4 mentions)
Gentamicin, by Thermo Fisher Scientific (4 mentions)
Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (4 mentions)
Rpmi 1640, by Thermo Fisher Scientific (4 mentions)
Horse serum, by Thermo Fisher Scientific (4 mentions)
Mda mb 231, by American Type Culture Collection (4 mentions)
Penicillin g, by Thermo Fisher Scientific (4 mentions)
Polyjet in vitro dna transfection reagent, by SignaGen (3 mentions)
99 protocols using dmem ham s f12 medium
1
KGN Cell Culture and Stimulation
2
Bladder Cancer Cell Lines Cultivation
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The human bladder cancer cell lines RT4, T24, T24T, and UMUC3 and the normal urinary epithelial cell lines SV-HUC-1 were used in the study. All cancer cell lines were generously provided by Dr. Haishan Huang (Wenzhou Medical University, Zhejiang, China), and they were subjected to DNA tests and authenticated before use for the studies. UMUC3 cells were maintained at 37°C in a 5% CO2 incubator in DMEM (Gibco, Grand Island, NY, USA), supplemented with 10% fetal bovine serum (FBS) (Gibco, Grand Island, NY, USA). T24 and T24T cells were cultured with a 1:1 mixture of DMEM-Ham’s F12 medium (Gibco, Grand Island, NY, USA), supplemented with 5% FBS. RT4 cells were cultured with 1640 (Gibco, Grand Island, NY, USA), supplemented with 10% FBS. SV-HUC-1 cells were cultured with F-12K medium supplemented with 10% FBS. Stable transfections were performed with constructs using PolyJet DNA In Vitro Transfection Reagent (SignaGen Laboratories, Gaithersburg, MD, USA), according to the manufacturer’s instructions, and stable transfectants were selected with puromycin (0.2–0.3 mg/mL) or hygromycin B (200–400 mg/mL) for 3 or 4 weeks according to the different antibiotic resistance plasmids transfected.42 (link), 50 (link)
3
Culturing C3A and iCSCs Cells
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All cells were cultured at 37°C with 5% CO2. C3A cells were derived from the human hepatoma cell line HepG2. The culture medium for C3A cells was Eagle's minimum essential medium (Gibco, New York) containing 10% fetal bovine serum (HyClone, Logan), 0.1 mM non-essential amino acids (Gibco, New York), and 0.1 mM 2-mercaptoethanol (Gibco, New York). Culture medium for iCSCs cells was Dulbecco's modified Eagle's medium (DMEM)/Ham's F-12 medium (Gibco, New York) containing 20% knockout serum replacement (Gibco, New York), 1 mM L-glutamine, 0.1 mM nonessential amino acids, 0.1 mM 2-mercaptoethanol, and 10 ng/ml recombinant human basic fibroblast growth factor (bFGF; Life Technologies, New York). Suspension culture medium was iCSCs medium without bFGF. All iCSCs cells were maintained on Matrigel (BD Biosciences, Franklin Lakes) and passaged with 0.5 mM EDTA.
4
Oligodendrocyte Differentiation from hiPSC-derived NPCs
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A human NPC line, established from human-induced pluripotent stem cells (hiPSCs), was obtained from Royan Institute, Tehran, Iran [23 (link)], and used. NPCs were passaged in a 1:3 ratio for expansion on poly-d -lysine (PDL)-coated plates and cultured in neurobasal medium (Gibco) supplemented with 1× penicillin/streptomycin, 25 ng/ml bFGF, 20 ng/ml epidermal growth factor (EGF), and 2 mM L-glutamine (all from Invitrogen).
At about 70% confluency, OPC differentiation was induced according to a previously published protocol with minor modifications [24 ]. Briefly, NPCs were grown for 3 weeks in the oligo medium containing serum-free DMEM/HAMS F12 medium (Gibco) supplemented with 1% bovine serum albumin, 2 mM L-glutamine, 50 μg/ml gentamicin, 1× N2 supplement, 3 nM T3 (SIGMA), 2 ng/mL Shh (SIGMA), 2 ng/mL NT-3 (SIGMA), 20 ng/mL bFGF, and 10 ng/mL PDGF-AA (SIGMA). Differentiation of OPCs to OLs was initiated by growth factors withdrawn for 2 days.
Human embryonic kidney cells (HEK293T) were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS, Hyclone, USA) and 1% antibiotics (100 U/ml penicillin and 100 mg/ml streptomycin sulfate). Cells were grown in a humidified atmosphere containing 5% CO2 at 37 °C.
At about 70% confluency, OPC differentiation was induced according to a previously published protocol with minor modifications [24 ]. Briefly, NPCs were grown for 3 weeks in the oligo medium containing serum-free DMEM/HAMS F12 medium (Gibco) supplemented with 1% bovine serum albumin, 2 mM L-glutamine, 50 μg/ml gentamicin, 1× N2 supplement, 3 nM T3 (SIGMA), 2 ng/mL Shh (SIGMA), 2 ng/mL NT-3 (SIGMA), 20 ng/mL bFGF, and 10 ng/mL PDGF-AA (SIGMA). Differentiation of OPCs to OLs was initiated by growth factors withdrawn for 2 days.
Human embryonic kidney cells (HEK293T) were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS, Hyclone, USA) and 1% antibiotics (100 U/ml penicillin and 100 mg/ml streptomycin sulfate). Cells were grown in a humidified atmosphere containing 5% CO2 at 37 °C.
5
Differentiation of SH-SY5Y Neuroblastoma Cells
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SH-SY5Y cells (ATCC® CRL-2266) were grown in DMEM/HAM’s F12 medium (Gibco, Waltham, MA, USA) supplemented with 10% FBS until reaching confluency. Cell differentiation was initiated by the addition of 10 μM retinoic acid dissolved in cell culture medium containing 0.5% dimethyl-sulfoxide and 2% FBS for five days [28 (link)]. For experiments, only differentiated cultures of SH-SY5Y were used.
6
Cell Line Cultivation and Transfection
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Normal bronchus epithelial cell line Beas-2B and lung cancer cell lines A549, H1975, HCC827, H1299, and SK-MES-1 were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA). All cells were subjected to DNA analysis and authenticated before use in these studies [26 (link)]. A549 and SK-MES-1 cells were cultured in F12K (21127-022; Gibco/ThermoFisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine serum (FBS) (10437-028; Gibco). H1975 cells were cultured in a 1:1 mixture of DMEM/Ham’s F12 medium (10565-018; Gibco) supplemented with 5% FBS. HCC827 and H1299 cells were cultured in RPMI 1640 (11875-093; Gibco) supplemented with 10% FBS, and Beas-2B cells were cultured in DMEM (11995-065; Gibco) supplemented with 10% FBS. Cell transfections were performed using PolyJet DNA In Vitro Transfection Reagent (SignaGen Laboratories, Rockville, MD, USA) according to the manufacturer's instructions. For stable cell line selection, cells were treated with G418 (500–1000 μg/mL) or puromycin (0.2–0.3 μg/mL) depending on the antibiotic resistance plasmid used. Cells surviving the antibiotic selection were pooled as stable mass transfectants [27 (link)].
7
Bladder Cancer Cell Lines Characterization
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The human BC cell lines RT4, T24, UMUC3, and TCCSUP and the normal urinary epithelial cell lines SV-HUC-1 and UROtsa were used in the study. All cancer cell lines and SV-HUC-1 were subjected to DNA tests and authenticated before use for the studies. UROtsa was generously provided by Dr. Scott H. Garrent (University of North Dakota, Grand Forks, ND, USA).23 (link), 41 (link) UMUC3 cells were maintained at 37°C in a 5% CO2 incubator in DMEM (No. 11995-065; Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS) (No. 10437-028; Gibco). T24 cells were cultured with a 1:1 mixture of DMEM/Ham’s F12 medium (No. 10565-018; Gibco) supplemented with 5% FBS. UROtsa and RT4 cells were cultured with 1640 (No. 11875-093; Gibco) supplemented with 10% FBS, and TCCSUP cells were cultured with MEM (No. 11095-080; Gibco) supplemented with 10% FBS. SV-HUC-1 cells were cultured with F-12K medium supplemented with 10% FBS (No. 21127-022; Gibco) medium. Stable transfections were performed with constructs using PolyJet DNA In Vitro Transfection Reagent (SignaGen Laboratories, Gaithersburg, MD, USA) according to the manufacturer’s instructions, and stable transfectants were selected with puromycin (0.2–0.3 μg/mL) or hygromycin B (200–400 μg/mL) for 3 or 4 weeks according to the different antibiotic resistance plasmids transfected.23 (link), 42 (link)
8
Cell Culture Protocols for Diverse Cell Lines
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Human Embryonic Kidney (HEK293T/17 ATCC® CRL-11268) were grown in Dulbecco's modified Eagle Medium (DMEM, Gibco, USA) with 10% (v/v) fetal calf serum (FCS, PAA, Argentina) and 2 mM glutamine.
Chinese Hamster Ovary (CHO-K1, ATCC® CCL-61) cells were cultured in DMEM/Ham's F12 medium (Gibco, USA), supplemented either with 5% (v/v) FCS (growth medium) or 0,5% (v/v) FCS (production medium).
Madine Darby Bovine Kidney (MDBK, ATCC® CCL-22) cells were grown in minimum essential medium (MEM; Gibco, USA) supplemented with 10% (v/v) FCS (growth medium). Antiviral activity assays were performed using MEM medium supplemented with 2% (v/v) FCS (assay medium).
Human Daudi cell line was obtained from the Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSMZ; Braunschweig, Germany) culture collection (DSMZ n°: ACC 78). Daudi cells and human Peripheral Blood Mononuclear Cells (PBMCs) were cultured in Roswell Park MEMorial Institute 1640 medium (RPMI, Gibco, USA) with 10% (v/v) FCS. The same medium was then used for bioassays.
All cell cultures were maintained at 37 °C in humidified 5% CO2.
Chinese Hamster Ovary (CHO-K1, ATCC® CCL-61) cells were cultured in DMEM/Ham's F12 medium (Gibco, USA), supplemented either with 5% (v/v) FCS (growth medium) or 0,5% (v/v) FCS (production medium).
Madine Darby Bovine Kidney (MDBK, ATCC® CCL-22) cells were grown in minimum essential medium (MEM; Gibco, USA) supplemented with 10% (v/v) FCS (growth medium). Antiviral activity assays were performed using MEM medium supplemented with 2% (v/v) FCS (assay medium).
Human Daudi cell line was obtained from the Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSMZ; Braunschweig, Germany) culture collection (DSMZ n°: ACC 78). Daudi cells and human Peripheral Blood Mononuclear Cells (PBMCs) were cultured in Roswell Park MEMorial Institute 1640 medium (RPMI, Gibco, USA) with 10% (v/v) FCS. The same medium was then used for bioassays.
All cell cultures were maintained at 37 °C in humidified 5% CO2.
9
3D Culture of TNBC and MCF-10A Cells
10
Establishing Breast Cancer Cell Lines
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Breast cancer cell lines MDA-MB-231, MDA-MB-468, HCC1937, BT20, HCC1143, BT549, and Hs578T cells were procured from American Type Culture Collection (ATCC). HCC70, T47D, ZR75, SUM149, CAL51, and SUM159 were obtained from A. Toker (Beth Israel Deaconess Medical Center, Boston, MA), 4T1, 67NR, 4T07, MCF7, and HEK293T cells from R. Weinberg (Whitehead Institute, Cambridge, MA), and MCF-10A from J. Brugge (Harvard Medical School, Boston, MA). Human primary breast cancer cells DT22 were a gift from D. El-Ashry (Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, Miami, FL)24 (link). Bone marrow-derived human mesenchymal stem cells (BM-MSCs) were purchased from the Institute for Regenerative Medicine at Scott and White, Texas A&M Health Science Center (Temple, TX). Established cancer cell lines were cultured according to ATCC recommendations. DT22 cells and BM-MSCs were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with 10%fetal bovine serum (FBS). MCF10A cells were cultured in DMEM/Ham’s F12 medium supplemented with 5% horse serum (Gibco BRL, Waltham, MA), insulin (10 μg/ml; Wisent Bioproducts, Saint-Jean Baptiste, QC), hydrocortisone (500 ng/ml; Sigma–Aldrich, St. Louis, MO), epidermal growth factor (EGF) (20 ng/ml; R&D, Minneapolis, MN), and cholera toxin (100 ng/ml; List Biological Laboratories, Campbell, CA).
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