Two different Activin A-derived primed hESC lines (U-11-4-A3; female and U-12-3-A3, male) [19 (link)] were maintained on a mitomycin-inactivated feeder layer of mouse embryonic fibroblasts (MEFs) in hESC medium (Gibco KnockOut-Dulbecco’s Modified Eagle Medium (KO-DMEM) (Thermo Fisher Scientific, Waltham, MA, USA), 20% Gibco KnockOut-serum replacement (KOSR) (Thermo Fisher Scientific), 1% Gibco Penicillin/Streptomycin (Thermo Fisher Scientific), 1% Gibco non-essential amino acids (NEAA) (Thermo Fisher Scientific), 0.4 mM Gibco L-Glutamine (Thermo Fisher Scientific), 0.1 mM Gibco β-mercaptoethanol (Thermo Fisher Scientific)) supplemented with 4 ng/mL FGF2 (Peprotech, Rocky Hill, NJ, USA) and 20 ng/mL Activin A (R&D Systems, Minneapolis, MN, USA). All cultures were maintained at 37 °C in hypoxic conditions (5% O2 and 6% CO2).
Gibco l glutamine
Manufactured by Thermo Fisher Scientific
Sourced in United States, Belgium
Gibco L-Glutamine is a critical amino acid that serves as an essential nutrient for cell growth and proliferation in cell culture applications. It provides a source of nitrogen and carbon to support various cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Gibco penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Hepes buffer, by Merck Group (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Gentamycin, by Merck Group (2 mentions)
Minimum essential medium (mem), by Lonza (2 mentions)
Gibco non essential amino acids, by Thermo Fisher Scientific (1 mentions)
Fibroblast growth factor 2 (fgf2), by Thermo Fisher Scientific (1 mentions)
Activin a, by Thermo Fisher Scientific (1 mentions)
Activin a, by R&D Systems (1 mentions)
Rotenone, by Merck Group (1 mentions)
Xf fluxpaks, by Agilent Technologies (1 mentions)
Sodium pyruvate, by Merck Group (1 mentions)
Ferric ammonium citrate, by Merck Group (1 mentions)
1 14c oleic acid, by PerkinElmer (1 mentions)
Antimycin a, by Merck Group (1 mentions)
Seahorse xf dmem medium, by Agilent Technologies (1 mentions)
Carbonyl cyanide 4 trifluoromethoxy phenylhydrazone fccp, by Merck Group (1 mentions)
Xf 24 well microplates, by Agilent Technologies (1 mentions)
Ex cell acf dho medium, by Merck Group (1 mentions)
Z wehd fmk, by R&D Systems (1 mentions)
7 protocols using gibco l glutamine
1
Activin A-Derived primed hESC Culture
2
Investigating Oleic Acid Metabolic Pathways
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OA, conjugated OA-albumin from bovine serum (OA-BSA), methylated OA (me-OA), methyl-β-cyclodextrin (MBCD), oligomycin, carbonyl cyanide-4-(trifluoromethoxy) phenylhydrazone (FCCP), rotenone, antimycin A, glucose, sodium pyruvate, and ferric ammonium citrate (FAC) were purchased from Sigma-Aldrich (St. Louis, MO). 12-hydroxy OA (12-OH OA) was purchased from Cayman Chemical (Ann Arbor, MI). Two-hydroxy OA sodium salt (2-OH OA [Na+]) was purchased from Avanti Lipids (Alabaster, AL). 1-14C-oleic acid (58.2 mCi/mmol) was purchased from Perkin-Elmer (Boston, MA). Seahorse XF DMEM medium, XF24-well microplates, and XF Flux Paks were purchased from Agilent Technologies (Santa Clara, CA). Gibco L-glutamine was purchased from Thermo Fisher Scientific (Waltham, MA).
3
Encapsulation of CHO Suspension Cells
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CHO suspension
cells (Public Health England) were cultured with
a protein-free medium (EX-CELL ACF DHO Medium, Sigma-Aldrich, Germany)
supplemented with 4 mMl -glutamine (Gibco l -glutamine,
Thermo Fisher, USA); 4 × 106 cells in 200 μL
medium were encapsulated into the water-in-oil droplets.
cells (Public Health England) were cultured with
a protein-free medium (EX-CELL ACF DHO Medium, Sigma-Aldrich, Germany)
supplemented with 4 mM
Thermo Fisher, USA); 4 × 106 cells in 200 μL
medium were encapsulated into the water-in-oil droplets.
4
Murine B Cell Survival Assay
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Primary, spleen-derived murine B cells were purified and cultured at 37°C and 7.5% CO 2 in complete medium, Iscove's modified Dulbecco's medium (IMDM) (Biochrom, Merck) supplemented with 10% heat-inactivated FCS, 1% Gibco Penicillin-Streptomycin [penicillin (100 U/ml)-streptomycin (100 g/ml)], and 2 mM Gibco l- glutamine (all from Thermo Scientific). The cells were incubated either in medium alone or in the presence of human recombinant BAFF (100 ng/ml) (hrBAFF, ImmunoTools) over a period of 4 days. The medium was exchanged every second day and supplemented with fresh hrBAFF. Cell viability was assessed with a fixable viability dye (eBioscience). Annexin V and propidium iodide staining was not applicable for the survival assays because of reported false positivity by catfish egg lectin (108) . For the viability analysis, the percentages were obtained from the ungated, viability dye-negative (healthy) cells. The following inhibitors were first titrated on B cells and then used in this study: necrostatin (Nec-1), polycaspase inhibitor QVD (Q-VD-PH), and caspase-1 inhibitor Z-WEHD-FMK (all from R&D Biosystems).
5
Generation and Maintenance of LCLs
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For the generation of LCLs, B cells were isolated from PBMCs by positive selection using the MACS technology with CD19 microbeads (Miltenyi), according to the manufacturer’s protocol. Isolated B cells were resuspended in R10 (RPMI 1640 + 10% FBS, 1% penicillin/streptomycin + 1% L-Glutamine Gibco, Thermo Fisher Scientific). EBV particles were added at a MOI of 0.1 to the cells, which were then incubated at 37 °C, 5% CO2 until EBV-transformed B cells grew out. Established LCLs were passaged 2–3 times per week. LCLs provided by R. Marsh were generated in a similar way, by infecting total PBMCs with EBV collected from the spent supernatant of the B95-8 marmoset cell line and the addition of cyclosporine A. All cell lines were routinely checked for mycoplasma contamination.
6
Diabetes Drug Formulation Evaluation
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CS, viscosity 9 cP was sourced from Primex Ehf (Siglufjordur, Iceland); sodium ALG, from Memphis (Cairo, Egypt); REP powder and marketed tablet (Repaglinide® 1 mg), from EIPICO (Ash Sharqiyah, Egypt); leucine (LEU), from Fluka production GmbH (Buchs, Switzerland); MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide), HEPES buffer, fetal calf serum, gentamycin, dimethyl sulfoxide, sodium lauryl sulfate (SLS), and streptozotocin (STZ), from Sigma-Aldrich Co., St Louis, MO, USA; Minimum Essential Medium (MEM), from Lonza (Verviers, Belgium); and L-glutamine GIBCO®, from Thermo Fisher Scientific (Waltham, MA, USA).
7
Synthesis and Characterization of Functional Polymers
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All solvents and reagents were of analytical or HPLC grade and purchased from Sigma Aldrich (UK) or Fisher Scientific (UK) unless otherwise stated. Deuterated solvents were from Sigma Aldrich (UK) or Cambridge Isotopes (UK). Monomers used in this study (Polyethylene glycol methyl ether (PEGMA-ME 188, Mn 188), polyethylene glycol ethyl ether (PEGMA-EE 246, Mn 246), and polyethylene glycol methyl ether (PEGMA-ME475, Mn 475) were purchased from Sigma Aldrich (UK) and purified before use by passing through a column filled with neutral alumina. 2,2-azobisisobutyronitrile (AIBN) was purchased from Sigma Aldrich (UK) then recrystallized from methanol. Pentaerythritol Tetra (3-mercaptopropionate) (PETMP) and 2butanone were used as received from Sigma Aldrich (UK). Risedronate sodium was kindly supplied by SPIC Pharma (India). Spectra/Por dialysis membrane, 12,000-14,000 molecular weight cutoff, was purchased from Spectrum Laboratories (Canada). MTT (3-(4,5dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide), HEPES buffer, gentamycin, dimethylsulfoxide and sodium lauryl sulphate (SLS) from Sigma-Aldrich (UK). Foetal calf serum, Minimum Essential Medium (MEM), from Lonza (Belgium); and L-glutamine GIBCO ® , were purchased from Thermo Fisher Scientific (USA).
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