AML12 mouse hepatocytes, 293T cells and HepG2 cells were purchased from ATCC. MDA-MB-231 and MCF7 cells were gifts from Donald McDonnell’s laboratory. All cell lines were cultured in DMEM High Glucose media (Sigma Cat #5796) without sodium pyruvate and were supplemented with 10% FBS (Gibco). Cells were treated with the medium-chain fatty acids sodium hexanoate (C6), sodium octanoate (C8) and sodium decanoate (C10) (Sigma) dissolved in phosphate buffered saline (PBS) and sodium dodecanoate (C12) dissolved in 10% ethanol in PBS. PBS was used as the vehicle control in all experiments. In experiments containing C12 (Fig. 1F ), all samples also contained final concentration of 0.1% ethanol added to the medium. Cells were counted using a Beckman Coulter Countess cell counter using trypan blue exclusion and seeded at the indicated densities. All experiments were done in complete DMEM with 10% FBS (referred to as complete media) with fatty acids or vehicle supplemented, unless otherwise noted.
Dmem high glucose media
Manufactured by Merck Group
Sourced in United States
DMEM high glucose media is a cell culture medium that provides high levels of glucose to support the growth and metabolism of various cell types in vitro. It is a widely used formulation in cell biology research and pharmaceutical applications.
Automatically generated - may contain errors
Lab products found in correlation
L glutamine, by Merck Group (6 mentions)
Penicillin, by Merck Group (4 mentions)
Streptomycin, by Merck Group (4 mentions)
Penicillin, by Thermo Fisher Scientific (4 mentions)
Rpmi 1640 medium, by Merck Group (3 mentions)
Fetal bovine serum (fbs), by Merck Group (3 mentions)
2 mercaptoethanol, by Merck Group (2 mentions)
Sodium pyruvate, by Merck Group (2 mentions)
H3.3 lys 27 to methionine, by Merck Group (2 mentions)
Rpmi medium, by Merck Group (2 mentions)
Streptomycin, by Thermo Fisher Scientific (2 mentions)
Non essential amino acids (neaa), by Merck Group (2 mentions)
Aml12 mouse hepatocytes, by American Type Culture Collection (1 mentions)
Sodium decanoate c10, by Merck Group (1 mentions)
293t cell, by American Type Culture Collection (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Hepg2 cell, by American Type Culture Collection (1 mentions)
Nih 3t3 flp in, by Thermo Fisher Scientific (1 mentions)
T rex 293, by Thermo Fisher Scientific (1 mentions)
Dmem high glucose, by Merck Group (1 mentions)
17 protocols using dmem high glucose media
1
Fatty Acid Treatments in Cell Culture
2
Cell Culture Protocols for Multiple Cell Lines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
T-Rex-293 (Invitrogen), IMCD3 Flp-In (IMCD3; American Type Culture Collection; gift of Peter Jackson), Phoenix A (PhA; Indiana University National Gene Vector Biorepository), and 293FT cells were cultured in DMEM high glucose (Sigma-Aldrich; supplemented with 10% cosmic serum, 0.05 mg/ml penicillin, 0.05 mg/ml streptomycin, and 4.5 mM glutamine). NIH 3T3 Flp-In (Thermo Fisher) cells were cultured in DMEM high glucose media (D5796; Sigma) with 10% Bovine calf serum (BCS; Sigma-Aldrich), 0.05 mg/ml penicillin, 0.05 mg/ml streptomycin, and 4.5 mM glutamine. 3T3-L1 cells (gift of Peter Michaely, UTSW) were cultured in DMEM high glucose (Sigma-Aldrich) media with 10% fetal bovine serum, 0.05 mg/ml penicillin, 0.05 mg/ml streptomycin, 4.5 mM glutamine, and 8 ng/ml biotin. Cell lines were tested negative for Mycoplasma using the Mycoplasma PCR Detection Kit (Genlantis). Transfection of plasmids was done with Polyfect (QIAGEN) or polyethylenimine (PEI) max. Stable cell lines were generated by retroviral infection or transfection. In many cases, stable lines were flow sorted and further selected for GFP. Control and Tulp3 ko MEFs were from embryonic day 13.5 embryos (Legue and Liem, 2019 (link)). Immortalized WT and Arl13b mutant MEFs were gifts from Tamara Caspary (Larkins et al., 2011 (link); Gigante et al., 2020 (link)).
3
Cell Line Authentication and Treatment
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The human normal hepatocyte cell line MIHA and the HCC cell line SNU475 were kindly gifted to us by Professor Suk Woo Nam (The Catholic University, Seoul, Korea) and the HCC cell line HepG2 was purchased from ATCC (Manassas, VA, USA). All cell lines used in this study were grown in Roswell Park Memorial Institute (RPMI)-1640 or Dulbecco’s modified Eagle’s medium (DMEM)-high glucose media (Sigma-Aldrich, St. Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS) (Hyclone, Logan, UT, USA) and 1% penicillin-streptomycin (Invitrogen, Carlsbad, CA, USA). The cells were incubated in humidified conditions with 5% CO2 at 37 °C. Mycoplasma contamination of all cell lines was tested using BioMycoX® Mycoplasma PCR Detection Kit (Cellsafe, Suwon, Korea) and short tandem repeat (STR) profiling was performed for authentication.
Valproic acid sodium salt (VPA) (Figure 1 A(i)), doxorubicin hydrochloride (DOX) (Figure 1 A(ii)), sodium butyrate, N-acetylcysteine (NAC), 3-methyladenine (3-MA), chlorpromazine (CPZ), methyl-β-cyclodextrin (MβCD), LY 294002 (LY), Hoechst 33258, and acridine orange hydrochloride hydrate (AO) were acquired from Sigma-Aldrich. 2′,7′-Dichlorodihydrofluorescein diacetate (H2DCFDA) was purchased from Molecular Probes™ (Eugene, OR, USA).
Valproic acid sodium salt (VPA) (
4
Culturing Primary and Tumor Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Primary cells were cultured in RPMI-1640 medium (Sigma-Aldrich, Taufkirchen, Germany), supplemented with 10% fetal calf serum (FCS), 2 mM l -glutamine, 100 U/l penicillin, 0.1 mg/ml streptomycin, 100 mM non-essential amino acids (all gibco®, Thermo Fisher Scientific, Karlsruhe, Germany), 1 mM sodium pyruvate and 50 mM 2-mercaptoethanol (both Sigma Aldrich). T110299 tumor cells had been isolated from tumors of genetically-engineered Ptf1a-Cre KrasG12D p53fl/R172H (KPC) mice and kindly provided by Prof. Siveke (West German Cancer Center (WTZ), University Hospital Essen). T110299 cells were cultured in DMEM high glucose media (Sigma-Aldrich), supplemented with 10% FCS, 2 mM l -glutamine, 100 U/l penicillin and 0.1 mg/ml streptomycin (all gibco®). All cells were kept in a humidified incubator at 37 °C and 5% CO2 and were regularly tested for mycoplasma contamination.
5
Culturing pediatric glioblastoma cell lines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A 13-year-old boy’s high-grade glioblastoma PBT24 cell line cells were donated by Prof. M. M. Alonso (University of Navarra, Spain) [43 (link)] for the study. A 3-year-old girl’s diffuse intrinsic pontine glioblastoma (DIPG) SF8628 cell line cells—harboring the histone H3.3 Lys 27-to-methionine (Sigma Aldrich, St. Louis, MO, USA)—were also studied [44 ,45 (link)]. The PBT24 cells were cultivated in Roswell Park Memorial Institute 1640 (RPMI), medium (Sigma Aldrich, St. Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS; Sigma Aldrich, St. Louis, MO, USA) containing 100 IU/mL of penicillin and 100 µg/mL of streptomycin (P/S; Sigma Aldrich, St. Louis, MO, USA). The SF8628 cells were cultivated in Dulbecco’s Modified Eagle Medium (DMEM)–High Glucose Media (Sigma Aldrich, St. Louis, MO, USA), supplemented with 10% fetal bovine serum (FBS; Sigma Aldrich, St. Louis, MO, USA) containing 100 IU/mL of penicillin and 100 µg/mL of streptomycin (P/S; Sigma Aldrich, St. Louis, MO, USA) and 2 mM L-Glutamine (Sigma Aldrich, St. Louis, MO, USA). Cells were incubated at 37 °C in a humidified 5% CO2 atmosphere.
6
Generation of Gpr161 Knockout NIH 3T3 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
NIH 3T3-FlpIn cells were authenticated by and purchased from Thermo Fisher Scientific. They have tested negative for Mycoplasma. The Gpr161−/− NIH 3T3 Flp-In cell line was a gift from Rajat Rohatgi23 (link). The cells were cultured in DMEM-high glucose media (D5796; Sigma) with 10% BCS (Sigma-Aldrich), 0.05 mg/ml penicillin, 0.05 mg/ml streptomycin, and 4.5 mM glutamine. Stable knockout cell lines were generated by retroviral infection with pBABE constructs having untagged wild type or mutant GPR161 inserts followed by antibiotic selection. Single or multiple amino acid mutations in full-length GPR161 were generated using Q5 site-directed mutagenesis kit (NEB).
7
Maintenance of Murine Melanoma Cell Line
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
B16-F10, murine skin melanoma cells from C57BL/6J mice, were purchased from ATCC® (ATCC® CRL-6475™) and maintained in DMEM high-glucose media (Sigma-Aldrich, 51435C) supplemented with 10% (v/v) of heat-inactivated Foetal Bovine Serum (FBS, Biowest, S1650) and 2 mM of L-glutamine (Sigma-Aldrich, G5792) at 37°C in a humidified atmosphere with 5% CO2. Subculturing was performed every 2 days.
8
In Vitro Assays for Mammary Cell Lines
9
Analyzing Caprin-2 Modulation in AVP Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HEK 293T/17 cells (ATCC, USA) were plated into 6-well plates at a density of 300,000 cells/well in DMEM high glucose media (Sigma–Aldrich) supplemented with 10% (vol/vol) FBS, 2 mM L-glutamine and 1× NEAA (Sigma–Aldrich). The following day media was changed and cells were co-transfected with the rat genomic AVP structural gene under the transcriptional control of the CMV promoter (derived from pSP72-VP [Chooi et al., 1994 (link)]) in combination with either: (a) control pRRL.sin.cppt.CMV.eGFP.wpre vector (‘eGFP’), (b) Caprin-2 overexpression vector pRRL.sin.cppt.CMV.rCaprin2.ires.eGFP.wpre (‘Caprin-2’), (c) Caprin-2 overexpression vector and control scrambled shRNA vector pRRL.sin.U6.Scr_shRNA.cppt.cmv.GFP.wpre (‘Ctrl’) and (d) Caprin-2 overexpression vector and Caprin-2 shRNA vector pRRL.sin.U6.Caprin-2_shRNA.cppt.CMV.GFP.wpre (‘Cap2 shRNA’). Transfection with 2 µg of each plasmid and Lipofectamine LTX reagent (Life Technologies) was performed according to the manufacturer protocol. 48 hr after transfection cells were lysed with TRIzol reagent (Life Technologies) and subjected to RNA extraction, as described above.
10
Vero E6 Cell Culture Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Normal monkey kidney cell line, Vero E6, was cultured into 75 cm2 flasks in DMEM-high glucose media (Sigma-Aldrich, Taufkirchen, Germany) supplemented with 4500 mg/L glucose, 2 mM L-glutamine, 1 mM sodium pyruvate, 10% fetal bovine serum (FBS), and antibiotics (penicillin 100 IU/mL, streptomycin 100 µg/mL). The cell culture was kept under standard culture conditions (37 °C, 95% humidified air, and 5% CO2).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!