D glucose

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D-Glucose is a monosaccharide that serves as a primary source of energy in many biological systems. It is an essential component in various laboratory experiments and analyses.

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D-MEM low glucose with phenol red (3 citations)

16 protocols using d glucose

Comano Spring Water's Impact on Skin Repair

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Each of the 6 skin specimens was used to harvest paired control and experimental samples. The control samples were cultured in DMEM with 1.0 g/l D-glucose (Biochrom, Ltd., Cambridge, UK), 10% fetal bovine serum (FBS), 1% penicillin (10,000 U/ml) and streptomycin (10 mg/ml), 1% gentamicin (all from Biowest) and 10 ml/l 200 mM L-glutamine (Eurobio Laboratoires, Les Ulis, France), and the central skin loss region was treated with a constant volume (200 µl) of sterile saline solution.
The samples treated with Comano spring water (treated samples) were cultured with DMEM powder without NaHCO₃ with 1.0 g/l D-glucose (Biochrom, Ltd.) and 10 ml/l 200 mM L-glutamine, reconstituted with filtered Comano spring water and enriched with 10% FBS, 1% penicillin (10,000 U/ml) and streptomycin (10 mg/ml) and 1% gentamicin (all from Biowest). The central skin loss region was treated with a constant volume (200 µl) of filtered Comano spring water. Following treatment, the control and treated samples were incubated at 37°C for 24, 48 and 72 h.

Nicoletti G., Saler M., Pellegatta T., Tresoldi M.M., Bonfanti V., Malovini A., Faga A, & Riva F. (2017). Ex vivo regenerative effects of a spring water. Biomedical Reports, 7(6), 508-514.

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Rat-Derived OLN93 Pre-OL Cell Line Protocol

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The OLN93 cell line was obtained from Prof. Dr. Christiane Richter-Landsberg (Universität Oldenburg, Germany). It is a rat derived (female), pre-OL adherent cell line derived from spontaneously transformed cells in primary rat brain glial cultures [50 (link)]. The cell line was authenticated by the Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany.
The cells were cultured as per the protocol mentioned by Gerstner et al. [51 (link)] in Dulbecco’s Modified Eagle’s Medium (DMEM) (with 3.7 g/l NaHCO₃, 25 mM HEPES, 4.5 g/l D-Glucose, 4.4 g/l NaCl; Biochrom), supplemented with 10% heat inactivated fetal calf serum (FCS, Biochrom), 0.01% human serum albumin (HSA, Grifols), and 1% penicillin-streptomycin solution. Cultures were kept in a 37 °C, 5% CO₂ incubator, and media was exchanged every 2–3 days.
B104 neuroblastoma cell line was purchased from the American Type Culture Collection (ATCC). The cells were used only for preparing conditioned media for OPC culture as described previously [43 ].

Sunny D.E., Hammer E., Strempel S., Joseph C., Manchanda H., Ittermann T., Hübner S., Weiss F.U., Völker U, & Heckmann M. (2020). Nup133 and ERα mediate the differential effects of hyperoxia-induced damage in male and female OPCs. Molecular and Cellular Pediatrics, 7, 10.

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Glioblastoma Cell Culture Protocol

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Two human glioblastoma cell lines were used. U-251 MG human brain glioblastoma cell line was obtained from CLS (Heidelberg, Germany) and U-373 MG was a generous gift from Dr. Bardenheuer (Essen, Germany). The cell lines U-251 and U-373 were routinely grown in Dulbecco’s modified eagle medium (DMEM) with 4.5 g/l d-Glucose, 3.7 g/l NaHCO₃, stable glutamine and Na-pyruvat (Biochrom AG, FG 0445, Germany). Media were supplemented with 10% sterile fetal bovine serum (Biochrom AG, S 0115) and 1% penicillin–streptomycin (Sigma-Aldrich, P0781). Cell lines were maintained at 37°C, 5% CO₂, and 90% humidity.

Krcek R., Matschke V., Theis V., Adamietz I.A., Bühler H, & Theiss C. (2017). Vascular Endothelial Growth Factor, Irradiation, and Axitinib Have Diverse Effects on Motility and Proliferation of Glioblastoma Multiforme Cells. Frontiers in Oncology, 7, 182.

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Microcontact Printing with Engineered NIH 3T3 Cells

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We used NIH 3T3 (ATCC CRL1658) fibroblasts for experiments involving microcontact printing. The NIH 3T3 cell culture was a gift from Louis Lim (Institute of Molecular and Cell Biology, ASTAR Singapore). For long-term experiments we employed a genetically modified variant termed NIH 3T3 X2 [39 (link)]. Compared with standard 3T3 cells, an enlarged fraction of X2 cells exhibited CDR formation. Cells were grown under standard conditions in Dulbecco’s MEM containing 3.7 g/L NaHCO₃, 4.5 g/L D-Glucose (Biochrom), 100 µg/ml Penicillin/Streptomycin (PAA), and 10% Fetal Bovine Serum (Biochrom). Cells were split at 80% confluency using Trypsin/EDTA (Biochrom). Cells were mycoplasma free.

Bernitt E., Koh C.G., Gov N, & Döbereiner H.G. (2015). Dynamics of Actin Waves on Patterned Substrates: A Quantitative Analysis of Circular Dorsal Ruffles. PLoS ONE, 10(1), e0115857.

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Engineered Human Cardiac Tissues from iPSCs

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EHTs were produced as previously explained [8 (link),16 (link)], and the experimental design is described in Figure 1B. Briefly, iPSC-CMs at day 14 were mechanically selected from the differentiation plate and digested with 5 mg/mL of collagenase type 1 (Worthington Biochemical Corporation, Lakewood, NJ, USA) for 45 min at 37 °C under agitation and later with 2 mg/mL of collagenase type 2 (Worthington Biochemical Corporation) for 45 min at 37 °C. Dissociated iPSC-CMs were counted (~1–2 × 10⁶ cells) and combined with 5 mg/mL of bovine fibrinogen (Merck, Darmstadt, Germany), 100 μL/mL of Matrigel (BD), DMEM (2 × 1 g/L D-Glucose, Biochrom), and 0.25 U/mL of thrombin (Merck), pipetted into the 2% agarose (Thermofisher Scientific) molds previously solidified in a 24-well culture dish with the silicone posts racks. After 90 min at 37 °C and 7% CO₂, 300 μL of the cell culture medium was added to easily remove the EHTs, and they were transferred into a new 24-well plate. EHTs were maintained at 37 °C in a 7% CO₂ humidified cell culture incubator with media changes three times a week and functional measurements twice a week. The EHT medium consisted of DMEM (Biochrom, Cambridge, UK), 10% inactivated horse serum (Merck), 1% penicillin/streptomycin (Merck), 10 μg/mL of insulin (Merck), and 33 μg/mL of aprotinin (Merck).

Llucià-Valldeperas A., Smal R., Bekedam F.T., Cé M., Pan X., Manz X.D., Wijnker P.J., Vonk-Noordegraaf A., Bogaard H.J., Goumans M.J, & de Man F.S. (2021). Development of a 3-Dimensional Model to Study Right Heart Dysfunction in Pulmonary Arterial Hypertension: First Observations. Cells, 10(12), 3595.

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Fibroblast Live Cell Imaging Assay

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NIH 3T3 (ATCC CRL1658) and NIH 3T3 X2 (ref. 54 (link)) fibroblasts were grown under standard conditions in Dulbecco's MEM containing 3.7 NaHCO₃, 4.5 D-Glucose (Biochrom), 100 Penicillin/Streptomycin (PAA), and 10% fetal bovine serum (Biochrom). Cells were mycoplasm free.
Live cell imaging was carried out on a Zeiss Axio Oberver.Z1 equipped with an incubation system at 37 °C and 5% CO₂. A Zeiss Achro Plan 10 × (NA 0.25) and Zeiss Plan Apochromat 40 × (NA 0.95) were used for phase contrast and fluorescence imaging respectively in conjunction with a Zeiss AxioCam MRm camera. No PDGF was added to the cell medium in experiments with live cell imaging (except for Supplementary Fig. 5). Actin was visualized using pLifeAct-TagGFP2 (Ibidi) and Lipofectamin2000 (Invitrogen) transfection.
Confocal imaging of fixed cells was done using a Zeiss LSM 780 equipped with a Plan-Apochromat 63 × (NA 1.4) objective. Actin was stained via Rhodamin/Phalloidin (Biotium) and the nucleus with DAPI (Roche). CDRs initiated via 30 hPDGF-BB (Cell Signaling Technology) in serum-free DMEM. Cells were then fixed 20 min after stimulation using methanol/acetate (1:1).

Bernitt E., Döbereiner H.G., Gov N.S, & Yochelis A. (2017). Fronts and waves of actin polymerization in a bistability-based mechanism of circular dorsal ruffles. Nature Communications, 8, 15863.

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Cultivation and Treatment of Glioblastoma Cells

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U-87 MG was obtained from the American Type Culture Collection (ATCC) (Manassas, USA) and U-251 MG from CLS (Heidelberg, Germany). Cells were cultivated in Dulbecco's modified eagle medium (DMEM) containing 4.5 g/L D-Glucose, 3.7 g/L NaHCO3, stable glutamine and Na-Pyruvat (Biochrom AG, FG 0445, Germany) and supplemented with 10% sterile fetal bovine serum (Biochrom AG, S 0115) and 1% penicillin (Sigma-Aldrich). Cells were maintained at 37°C, 5% CO2 and 90% air moisture. Subcultures were made on 12 mm Ø cover slips to apply VEGF, axitinib or radiation on the confluent cell monolayer.

Krcek R., Latzer P., Adamietz I.A., Bühler H, & Theiss C. (2017). Influence of vascular endothelial growth factor and radiation on gap junctional intercellular communication in glioblastoma multiforme cell lines. Neural Regeneration Research, 12(11), 1816-1822.

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Hormone Receptor-Positive Breast Cancer Cell Culture

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MCF-7 cells were chosen for their hormone receptor-positive attributes, reflecting the luminal breast cancer subtype. Their estrogen and progesterone receptor presence aligns with our hormone-related focus. Our selection was guided by their relevance to our objectives, well-documented status, and specific interest in MTE's impact on this hormone receptor-positive subtype. Additionally, our choice of MCF-7 cells was influenced by the existing data on green tea phenol's interaction with ERα, while information about ERβ interactions remains limited.
MCF-7 cells were cultured on an 80% monolayer in a cell culture bottle. Dulbecco's modified Eagle medium (DMEM; 3.7 g/l NaHCO3, 4.5 g/l d-glucose, 1.028 g/l stable glutamine, and sodium pyruvate; Biochrom, Berlin, Germany) supplemented with 10% heat-inactivated fetal calf serum (FCS; Biochrom, Berlin, Germany) was used for cultivation. The cells were incubated with atmospheric concentrations of CO₂ of 5% at 37 °C and were then trypsinized and counted for further use.

Keckstein S., Tilgener C., Jeschke U., Hofmann S., Vilsmaier T., Keilmann L., Heidegger H., Kaltofen T., Batz F., Mahner S, & Schröder L. (2023). Effects of matcha tea extract on cell viability and estrogen receptor-β expression on MCF-7 breast cancer cells. Archives of Gynecology and Obstetrics, 309(4), 1509-1514.

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Phage Cytotoxicity Assay in A549 Cells

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The cell line A549 (ATCC CCL-185) was grown in Dulbecco’s Modified Eagle’s Medium (DMEM with 3.7 g/L NaHCO₃ and 4.5 g/L D-glucose, Biochrom) supplemented with 10% of fetal bovine serum (FBS, Biochrom) and 1% Penicillin/Streptomycin (Sigma-Aldrich), at 37 °C in a humidified incubator with 5% CO₂. Cells were seeded into wells of a 96-well plate at 1 × 10⁶ cells/mL and incubated for 24 h, to reach a cell confluence between 70 and 90%. Phage solution (in SM buffer) was added to the cells, so that a phage concentration of 10⁸ PFU/mL and a final volume of 100 μL were achieved in the wells. Controls were performed by adding the same amount of SM buffer to the wells with cells grown in the same conditions. After 24 h, 20 μL of CellTiter 96 Aqueous One Solution Reagent (Promega) was added to each well and incubated for 1 h at 37 °C in a humidified 5% CO₂ atmosphere, protected from light. The amount of soluble formazan produced by cellular reduction of MTS was measured by reading the absorbance at 490 nm. The experiment was performed in triplicate.

Oliveira H., Pinto G., Oliveira A., Noben J.P., Hendrix H., Lavigne R., Łobocka M., Kropinski A.M, & Azeredo J. (2017). Characterization and genomic analyses of two newly isolated Morganella phages define distant members among Tevenvirinae and Autographivirinae subfamilies. Scientific Reports, 7, 46157.

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Trophoblast Cell Line Stimulation Assay

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The choriocarcinoma cell lines BeWo (ECACC, Salisbury, UK) and JEG-3 (ATCC), which are useful models of human trophoblasts, were used for the study. The cells were cultured in DMEM (3.7 g/L NaHCO₃, 4.5 g/L d-glucose, 1.028 g/L stable glutamine, and Na-Pyruvate; Biochrom, Berlin, Germany). 10% heat-inactivated FCS was added to the medium, and the solution was incubated at an atmospheric CO₂ concentration of 5% and at 37°C.
BeWo and JEG-3 cells were separately grown on sterile 12 multiwell slides at a density of 500,000 cells/mL DMEM with 10% foetal cow serum. After 4 h, medium was changed to pure DMEM. The cells were stimulated with 0.01 nM or 0.1 nM triiodothyronine (T3) (Sigma T2877; Lot:106K1157V, Sigma-Aldrich), thyronamine (T₀AM) (Fluka 80345, Lot: BCBL2185V, Buchs, Switzerland), 3-Iodothyronamine (T₁AM) (Cayman Chemical) and RO5203548 (Glixx Laboratories, Hopkinton, MA, USA) for 2 h (PCR samples) and 48 h (Immunocytochemistry and Western blot samples), respectively. Control cells were incubated without stimulants. As a control solvent, the following buffers were used: The stimulant T3 has been diluted in 1 M NaOH in DMEM (Dulbecco’s Modified Eagle Medium) and T₁AM and RO5203548 were diluted in DMSO (Dimethylsulfoxide).

Stavrou S., Gratz M., Tremmel E., Kuhn C., Hofmann S., Heidegger H., Peryanova M., Hermelink K., Hutter S., Toth B., Mayr D., Mahner S., Jeschke U, & Vattai A. (2018). TAAR1 induces a disturbed GSK3β phosphorylation in recurrent miscarriages through the ODC. Endocrine Connections, 7(2), 372-384.

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