HeLa and HEK293T cells were grown at 37 °C in a 5.0% CO2 water vapour saturated incubator in DMEM growth medium (Gibco, Carlsbad, CA, USA), supplemented with 10% fetal bovine serum (Sigma, St. Louis, MO, USA).
Dmem growth medium
Manufactured by Thermo Fisher Scientific
Sourced in United States
DMEM (Dulbecco's Modified Eagle's Medium) is a widely used cell culture medium formulated to support the growth and maintenance of a variety of cell lines. It provides essential nutrients, amino acids, vitamins, and other components necessary for cell proliferation and survival in in vitro cell culture systems.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (11 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (10 mentions)
Trypsin edta, by Thermo Fisher Scientific (6 mentions)
L glutamine, by Thermo Fisher Scientific (4 mentions)
Penicillin, by Thermo Fisher Scientific (3 mentions)
Streptomycin, by Thermo Fisher Scientific (3 mentions)
Non essential amino acids (neaa), by Thermo Fisher Scientific (3 mentions)
Pen strep, by Thermo Fisher Scientific (3 mentions)
Tissue culture plastic, by Greiner (2 mentions)
Rpmi 1640 growth medium, by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Merck Group (2 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Rapamycin, by Merck Group (1 mentions)
Bleomycin, by Nippon Kayaku (1 mentions)
A549 human lung adenocarcinoma cell line, by American Type Culture Collection (1 mentions)
Fluvastatin sodium, by Selleck Chemicals (1 mentions)
Erlotinib hydrochloride, by Selleck Chemicals (1 mentions)
Nci h1993, by American Type Culture Collection (1 mentions)
Pitavastatin calcium, by Selleck Chemicals (1 mentions)
Calu 6, by American Type Culture Collection (1 mentions)
29 protocols using dmem growth medium
1
Cell Culture Conditions for HeLa and HEK293T
2
Lens Epithelial Cell Culture Protocol
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Human SRA01/04 lens epithelial cell (LEC) were cultured in high-glucose DMEM growth medium (Gibco, Thermo Fisher Scientific, USA) containing 10% FBS (Biowest, FRA), 1% penicillin (100µg/ml) and 1% streptomycin (100µg/ml) at 37°C in a 5% CO2 atmosphere. SRA01/04 cell line was authenticated by short tandem repeat (STR) analysis at Sun Yat-Sen University (Supplementary Figure S1 ). Rabbit primary lens epithelial cells were isolated from 7~8 weeks old New Zealand white rabbits and cultured in DMEM/F12 with 20% FBS, 1% penicillin, 1% streptomycin and 1% NEAA (Gibco, Life Technologies Corporation). All cell lines were negative for mycoplasma. Recombinant human TGFβ2 was purchased from Novoprotein. For TGFβ2 and THZ1 (Sellcek, CN) treatment, the cells were cultured in fresh culture medium with 10ng/ml recombinant human TGFβ2 and different concentrations of THZ1 for 24h.
3
Production and Characterization of Influenza DI Particles
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T25 flasks were seeded with a coculture of 293T cells stably expressing PB1, PB2, PA (1.4 × 106 cells) and MDCK cells stably expressing PB2 and PA (0.4 × 106 cells) in DMEM growth medium (Gibco). For the production of DIPs encoding two DI segments derived from IAV genomic segments 1 and 3 (S1S3 DIPs), cells were cotransfected with plasmids encoding DI RNA derived from segments 1 and 3 of PR8 origin, jointly with plasmids encoding IAV genomic segments 2 and 4 to 8 of WSN origin using the calcium phosphate method. Similarly, for production of DIPs expressing single DI segments with a deletion in segment 1 (S1 DIPs) or segment 3 (S3 DIPs), cells were cotransfected with 7 IAV genomic segments and either one DI segment derived from S1 (for S1 DIPs) or one derived from S3 (for S3 DIPs). After overnight incubation, cells were washed once with PBS and fresh DMEM infection medium (2% FBS, 1% pen/strep, without trypsin) was added. As negative control, parental MDCK and 293T cells were also transfected. Supernatants were harvested at 3, 5 and 7 days post transfection, cleared by centrifugation at 1500 × g for 10 min and stored at − 80 °C for further use. Titers for DIP supernatants were determined by focus formation assay on PB2, PA expressing MDCK cells, as described15 (link).
4
Bleomycin-Induced Lung Cancer Cell Assay
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The A549 human lung adenocarcinoma cell line was purchased from the American Type
Culture Collection (Manassas, VA, USA). The cells were cultured in DMEM growth
medium (Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum
(Gibco) and antibiotics (100 units/mL penicillin and 100 mg/mL streptomycin) at
37°C in a humidified air atmosphere containing 5% CO2. Bleomycin was
purchased from Nippon Kayaku Co. Ltd. (Tokyo, Japan), dissolved in 0.9%
phosphate-buffered saline (PBS), and stored at −20°C. A549 cells were incubated
with Bleomycin (0, 0.1, 1, 5, 10 and 50 µg/mL) for 120 hours in culture
medium.20 (link) The same volume of PBS was added to the culture medium
as a negative control. The mTOR inhibitor rapamycin was purchased from Sigma
(St. Louis, MO, USA), dissolved in dimethylsulfoxide (DMSO) (Sigma), and stored
at 4°C. The cells were treated with 100 mM rapamycin for 24 hour in the presence
or absence of Bleomycin. The same volume of DMSO was added to the culture medium
as a positive control.
Culture Collection (Manassas, VA, USA). The cells were cultured in DMEM growth
medium (Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum
(Gibco) and antibiotics (100 units/mL penicillin and 100 mg/mL streptomycin) at
37°C in a humidified air atmosphere containing 5% CO2. Bleomycin was
purchased from Nippon Kayaku Co. Ltd. (Tokyo, Japan), dissolved in 0.9%
phosphate-buffered saline (PBS), and stored at −20°C. A549 cells were incubated
with Bleomycin (0, 0.1, 1, 5, 10 and 50 µg/mL) for 120 hours in culture
medium.20 (link) The same volume of PBS was added to the culture medium
as a negative control. The mTOR inhibitor rapamycin was purchased from Sigma
(St. Louis, MO, USA), dissolved in dimethylsulfoxide (DMSO) (Sigma), and stored
at 4°C. The cells were treated with 100 mM rapamycin for 24 hour in the presence
or absence of Bleomycin. The same volume of DMSO was added to the culture medium
as a positive control.
5
Statins Overcome Erlotinib Resistance in NSCLC
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Experiments were carried out with human lung adenocarcinoma cell lines A549 (ATCC CCL_185), Calu6 (ATCC HTB-56) and NCI-H1993 (ATCC CRL-5909). All cell lines were obtained from American Type Cell Culture Collection (ATCC) and cultured in DMEM growth medium (GIBCO #31966-21), supplemented with 10% heat-inactivated foetal bovine serum (FBS) and 1% antibiotics (penicillin, streptomycin). Cells were kept at 37 °C and 5% CO2 in the incubator and were passaged at 80–90% confluence every 2–3 days to maintain continuous logarithmic growth. All cell lines studied in this work were erlotinib resistant and EGFR wild type (Table 1 ). Cells were treated with pitavastatin calcium (SelleckChem, #S1759), fluvastatin sodium (SelleckChem, #S1909) and erlotinib hydrochloride (SelleckChem, #S1023).
Human NSCLC cell lines harbouring different genetic mutations examined in the study.
| Cell line | EGFR | K-Ras | Met | p53 | Erlotinib |
|---|---|---|---|---|---|
| A549 | Wildtype | Mutated | Wildtype | Wildtype | Resistant |
| Calu6 | Wildtype | Mutated | Wildtype | Mutated | Resistant |
| H1993 | Wildtype | Wildtype | Amplified | Mutated | Resistant |
6
Lentiviral Production of Siglec-6
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Siglec-6 lentivirus was produced as previously reported by Bhattacherjee, A. et al.72 (link). Briefly, 1 × 106 HEK293T cells were plated in a 6-well dish containing 1.5 mL of DMEM growth medium (Gibco) containing 10% (V/V) fetal bovine serum (FBS; Gibco), 100 U/mL penicillin (Gibco) and 100 µg/mL streptomycin (Gibco). 24 h later, a mixture of 625 ng RP18, 625 ng RP19, 1250 ng hSiglec-6 vector, 7.5 µL TransIT®-LT1 Reagent (Mirus Bio), and Opti-MEM media (Gibco) was added to the HEK293T cells. Cells were incubated with this transfection mixture at 37 °C, 5% CO2 for 72 h. Following transfection, the cell supernatant was harvested and concentrated using Lenti-X concentrator reagent (Takara Bio) following the manufacturer’s instructions.
7
Cell Culture and Differentiation Protocols
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MDA-MB-231 cells were cultured in DMEM growth medium (Gibco, 11039-021) supplemented with 10% heat-inactivated fetal bovine serum (FBS) (Corning, MT35010CV) and 1× Anti-Anti (Thermo Fisher Scientific, 15240112) at 37°C with 5% CO2. Undifferentiated HC11 cells were cultured in growing medium (RPMI-1640; Thermo Fisher Scientific, 72400047), supplemented with 10% FBS, 5 µg/ml insulin (Millipore-Sigma, I6634), 10 ng/ml epidermal growth factor (EGF; Preprotech, AF-100-15), 1× Anti-Anti at 37°C with 5% CO2. Competent HC11 cells were primed for differentiation by culturing them in priming medium [RPMI-1640 supplemented with 5% charcoal-stripped-FBS (Equitech Bio, SFBM31), 5 µg/ml insulin, 1 µM dexamethasone (Millipore-Sigma, D4902-1G) and 1× Anti-Anti] for 18 h at 37°C with 5% CO2. To induce differentiation, primed HC11 cells were cultured in DIP Medium [RPMI-1640, supplemented with 10% FBS, 5 µg/ml insulin, 1 µM dexamethasone, 1× Anti-Anti and 3 µg/ml Prolactin (NHPP, oPRL-21)] at 37°C with 5% CO2. Primary cells were harvested from 8- to 12-week-old mice and cultured as previously described (Macias et al., 2011 (link); Rubio et al., 2020 (link)).
8
Cell Culture Protocol for J-Lat, 5A8, and U2OS
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The J-Lat 10.6 and 5A8 cells were cultured in flasks (Thermo Fisher Nunc™ EasYFlask™) in RPMI 1640 growth medium (L-glutamine, Gibco) with 10% fetal bovine serum (Gibco) and 1% penicillin-streptomycin (Gibco) and the human bone osteosarcoma epithelial (U2OS) cells were cultured as adherent cells directly on tissue culture plastic (Greiner) in DMEM growth medium (high glucose, L-glutamine, no sodium pyruvate; Gibco) with 10% fetal bovine serum (Gibco) and 1% penicillin-streptomycin (Gibco) at 37°C and 5% CO2. U2OS cells were harvested using 0.05% Trypsin-EDTA (Gibco).
9
Embryonic Cell Isolation and Culture
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E13.5-E15.5 bodies were collected from control, RoninF80L/F80L, and Hcfc1A115V/Y embryos and the livers and hearts were removed. The remaining tissue was transferred into a clean petri dish, and 300 µL of 0.05% trypsin-EDTA was added (Gibco, #25200056). The tissue was then minced using a clean razor blade for 5 to 7 min until single cells were released. The sample was then pipetted into a 15-mL conical tube containing 5 mL DMEM growth medium (Gibco, #11995065) with 10% fetal bovine serum (FBS) (Gibco, #26140079) and 1% penicillin/streptomycin (Pen/Strep) (Gibco, #15140122). After allowing the tissue to settle for 2 min, the supernatant with single cells was transferred to another clean 15-mL conical tube and centrifuged at room temperature (150 × g, 5 min). The media was removed, and the cell pellet was resuspended with 10 mL fresh DMEM. Cell suspension was then seeded into a clean 10-cm cell culture dish and incubated at 37 °C with 5% CO2 overnight before changing media and cell expansion.
10
Culturing Human Endometrial Cancer Cell Lines
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EC cell lines (Ishikawa and HEC1A) were obtained, authenticated and tested for Mycoplasma prior to usage. Cell lines were cultured in DMEM growth medium (Gibco Life Technologies; Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% (v/v) heat inactivated fetal bovine serum (Hi-FBS) (Gibco Life Technologies) and 2 mM glutamine (GlutaMAX). Cells were grown in 25 cm2 or 75 cm2 culture flasks (Corning, Tewksbury, MA, USA) and maintained at 37 °C, 95% humidity and 5% (v/v) CO2 (Thermo Fisher, Waltham, MA, USA). At 80% confluence, both cell lines were harvested, and a cell count conducted using a haemocytometer. Subculture was conducted multiple times and harvested cells pooled and pelleted by centrifugation at 1500× g for five minutes and stored at −80 degrees pending further analyses.
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