2-NBDG
(Synonyms: 2-(N-7-硝基-2,1,3-苯并恶二唑-4-氨基)-2-脱氧-D-葡萄糖) 目录号 : GC102892-NBDG是一种荧光标记的2-脱氧葡萄糖类似物,可用作细胞葡萄糖代谢评估的示踪剂(激发/发射波长:475/550纳米)。
Cas No.:186689-07-6
Sample solution is provided at 25 µL, 10mM.
- Ecotox Environ Safe 243 (2022): 113996.PMID:36030680
- Bioorganic Chemistry (2023): 106341.
- Oncogene (2023): 1-11.
- J Cardiovasc Dev Dis (2023): 10(6) 246.
- Front Pharmacol 13 (2022): 875014.PMID:35694255
- Ind Eng Chem Res (2023).
- Phytomedicine 114 (2023): 154740.PMID:36965373
- Nat Commun 15.1 (2024):557.PMID:38228638
- Tradit Med Res 9.7 (2024):37.
- Endocrinol Diab Meta 7.3 (2024).PMID:38556697
- Mol Carcinogen (2024).PMID:38634741
- A thesis of McGill University (2024).
- Cns Neurosci Ther 30.6 (2024).PMID:38887182
- bioRxiv (2024):2024-06.
- Environ Sci Technol (2024).PMID:39167196
- Endocrin Diab Metab 7.3 (2024):e00482.PMID:38556697
2-NBDG is a fluorescence-labeled 2-deoxy-glucose analog useful as a tracer for evaluation of cellular glucose metabolism (Ex/Em: 475/550 nm).
Glucose is a necessary source of energy for sustaining cell activities and homeostasis in tissues. Glucose metabolism is an important target in many diseases and changed with the pathological condition, therefore, evaluation of glucose metabolism can be a significant indication in disease progressions.
2-NBDG can be used in many kinds of cells in vitro, such as HepG2 human hepatocarcinoma cells, L6 rat skeletal muscle cells, MCF-7 breast cancer epithelial cells and astrocytes, it is also used in disease models, epilepsy rat, hyperglycemia, diabetes or mouse xenograft model of cancer.
2-NBDG enters cells through glucose transporters and is subsequently phosphorylated by hexokinase and trapped inside cells. Flow cytometric detection of fluorescence produced by cells can be performed to examine 2-NBDG uptake into living cells, and the intracellular concentration of transported 2-NBDG can be measured with a fluorescence microplate assay. It can be detected with a fluorescence imaging microscopy or CCD camera simply as well.
2-NBDG is a fluorescently labeled glucose tracer that is transported into cells by the same glucose transporter (GLUT) as glucose. Once 2-NBDG is taken up by cells, it is phosphorylated at the C-6 position to give 2-NBDG-6-phosphate, which is well retained in the cell. Compared to other glucose tracers such as 2-DG or FDG, 2-NBDG enables in situ measurement of 2-NBDG with high temporal and spatial resolution at the single-cell level. (suitable for fluorescence microscopy and flow cytometry detection)
Rationale for 2-NBDG glucose uptake assay in cells: Once 2 NBDG is taken up by cells, it is phosphorylated at the C-6 position to generate 2-NBDG-6-phosphate in 2 NBDG metabolism, which is well retained in the cell, the fluorescence intensity is proportional to the cellular glucose uptake activity.
References:
[1]. Zou C, Wang Y, Shen Z. 2-NBDG as a fluorescent indicator for direct glucose uptake measurement[J]. Journal of biochemical and biophysical methods, 2005, 64(3): 207-215.
[2]. O’Neil R G, Wu L, Mullani N. Uptake of a fluorescent deoxyglucose analog (2-NBDG) in tumor cells[J]. Molecular Imaging and Biology, 2005, 7(6): 388-392.
[3]. Tsytsarev V, Maslov K I, Yao J, et al. In vivo imaging of epileptic activity using 2-NBDG, a fluorescent deoxyglucose analog[J]. Journal of neuroscience methods, 2012, 203(1): 136-140.
[4]. Yan Chen, Junjian Zhang, Xiang-yang Zhang, 2-NBDG as a Marker for Detecting Glucose Uptake in Reactive Astrocytes Exposed to Oxygen-Glucose Deprivation In Vitro. J Mol Neurosci (2015) 55:126–130.
[5]. Tsytsarev V, Maslov K I, Yao J, et al. In vivo imaging of epileptic activity using 2-NBDG, a fluorescent deoxyglucose analog[J]. Journal of neuroscience methods, 2012, 203(1): 136-140.
2-NBDG是一种荧光标记的2-脱氧葡萄糖类似物,可用作细胞葡萄糖代谢评估的示踪剂(激发/发射波长:475/550纳米)。
葡萄糖是维持组织细胞活动和稳态所必需的能量来源。葡萄糖代谢在许多疾病中都是一个重要的靶点,并且会随着病理情况而改变,因此评估葡萄糖代谢可以成为一种重要的指标来判断疾病进展情况。
2-NBDG可以在许多种体外细胞中使用,例如HepG2人类肝癌细胞、L6大鼠骨骼肌细胞、MCF-7乳腺癌上皮细胞和星形胶质细胞,它还用于疾病模型,如癫痫大鼠、高血糖、糖尿病或小鼠异种移植的癌症模型。
2-NBDG通过葡萄糖转运体进入细胞,并被己糖激酶磷酸化并困在细胞内。可以使用流式细胞术检测细胞产生的荧光来检查2-NBDG进入活细胞的情况,也可以使用荧光微孔板分析法测量转运的2-NBDG在细胞内的浓度。它还可以简单地用荧光成像显微镜或CCD相机进行检测。
2-NBDG是一种荧光标记的葡萄糖示踪剂,通过与葡萄糖相同的葡萄糖转运体(GLUT)进入细胞。一旦2-NBDG被细胞吸收,它会在C-6位置磷酸化成为2-NBDG-6-phosphate,并且能够很好地保留在细胞内。与其他葡萄糖示踪剂如2-DG或FDG相比,2-NBDG可以以高时间和空间分辨率在单个细胞水平上原位测量2-NBDG。(适用于荧光显微镜和流式细胞术检测)
2-NBDG葡萄糖摄取实验在细胞中的原理:一旦细胞吸收了2-NBDG,它会在C-6位置被磷酸化,生成2-NBDG-6-磷酸盐,在2 NBDG代谢中很好地保留在细胞内,荧光强度与细胞内葡萄糖摄取活性成正比。
Procedure for 2-NBDG uptake assay for MEFs [1]
1. Mouse embryonic fibroblasts (MEFs) are isolated from the embryos of C57BL/6 WT mouse (13.5 days).
2. Culture the MEF cells until reaching 80-90% confluence in 10 cm Petri dishes with DMEM growth medium in a humidified cell culture incubator (37 °C, 5% CO2).
Note: Don’t use MEFs beyond passage 3. MEFs usually become senescent at about passage 4 to 5.
3. Remove culture medium and wash cells one time with 10 ml 1x PBS.
4. Trypsinize cells using 4 ml of 0.05% trypsin-EDTA for 3 min at 37 °C.
Note: Use room temperature or pre-warmed 1x PBS from Step A3 to Step A9. Using chilled 1x PBS after Step A9.
5. Transfer cells to 15 ml polystyrene centrifuge tubes.
6. Harvest cells at 200 x g for 5 min by centrifugation.
7. Wash pelleted cells one time with 5 ml 1x PBS.
8. Count cells using a hemocytometer chamber.
9. Incubate 1 x 106 MEF cells in a 37 °C water bath for 2 h with 1 ml of PBS containing 100 μM 2-NBDG. Incubate the same number of MEFs in the water bath with 1 ml PBS without 2-NBDG as a negative control.
10. Pellet the cells at 200 x g for 5 min by centrifugation. After washing the cells with chilled 1x PBS, the cells are pelleted at 200 x g for 5 min by centrifugation.
11. Resuspend cells in 0.5 ml of ice-cold 1x PBS with 2% FBS.
Note: Always keep cells on ice after this step.
12. Filter cells through a 35 µm nylon mesh (the cell-strainer cap of the 5 ml round-bottom polystyrene tubes) to obtain a uniform single-cell suspension in a 5 ml tube.
13. Keep the samples on ice until analysis on a flow cytometer.
14. Perform flow cytometric analysis. Acquire 10,000 single-cell events per reaction.
15. Analyze fluorescence intensity
Procedure for 2-NBDG uptake assay for breast cancer cells [1]
Using the same procedure as MEFs’ uptake assay except incubating 1 x 106 MCF7 cells in a 37 °C water bath only for 30 min (instead of two hours for MEFs) with 1 ml of PBS containing 100 μM 2-NBDG.
10 mM stock of 2-NBDG: Dissolve 5 mg 2-NBDG in 1.46 ml PBS. Store at -20 °C in the dark
This protocol only provides a guideline, and should be modified according to your specific needs
References:
[1]. Dong, S., Baranwal, S., Garcia, A., Serrano-Gomez, S. J., Eastlack, S., Iwakuma, T., Mercante, D., Mauvais-Jarvis, F. and Alahari, S. K. (2017). Nischarin inhibition alters energy metabolism by activating AMP-activated protein kinase. J Biol Chem 292(41): 16833-16846.
Cas No. | 186689-07-6 | SDF | |
别名 | 2-(N-7-硝基-2,1,3-苯并恶二唑-4-氨基)-2-脱氧-D-葡萄糖 | ||
化学名 | (3R,4R,5S,6R)-6-(hydroxymethyl)-3-((7-nitrobenzo[c][1,2,5]oxadiazol-4-yl)amino)tetrahydro-2H-pyran-2,4,5-triol | ||
Canonical SMILES | OC[C@](O1)([H])[C@](O)([H])[C@@](O)([H])[C@](NC2=CC=C(N(=O)=O)C3=NON=C23)([H])C1([H])O | ||
分子式 | C12H14N4O8 | 分子量 | 342.26 |
溶解度 | ≥ 17.1mg/mL in Water with ultrasonic | 储存条件 | -20°C, protect from light |
General tips | 请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液;一旦配成溶液,请分装保存,避免反复冻融造成的产品失效。 储备液的保存方式和期限:-80°C 储存时,请在 6 个月内使用,-20°C 储存时,请在 1 个月内使用。 为了提高溶解度,请将管子加热至37℃,然后在超声波浴中震荡一段时间。 |
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Shipping Condition | 评估样品解决方案:配备蓝冰进行发货。所有其他可用尺寸:配备RT,或根据请求配备蓝冰。 |
制备储备液 | |||
1 mg | 5 mg | 10 mg | |
1 mM | 2.9218 mL | 14.6088 mL | 29.2176 mL |
5 mM | 0.5844 mL | 2.9218 mL | 5.8435 mL |
10 mM | 0.2922 mL | 1.4609 mL | 2.9218 mL |
第一步:请输入基本实验信息(考虑到实验过程中的损耗,建议多配一只动物的药量) | ||||||||||
给药剂量 | mg/kg | 动物平均体重 | g | 每只动物给药体积 | ul | 动物数量 | 只 | |||
第二步:请输入动物体内配方组成(配方适用于不溶于水的药物;不同批次药物配方比例不同,请联系GLPBIO为您提供正确的澄清溶液配方) | ||||||||||
% DMSO % % Tween 80 % saline | ||||||||||
计算重置 |
计算结果:
工作液浓度: mg/ml;
DMSO母液配制方法: mg 药物溶于 μL DMSO溶液(母液浓度 mg/mL,
体内配方配制方法:取 μL DMSO母液,加入 μL PEG300,混匀澄清后加入μL Tween 80,混匀澄清后加入 μL saline,混匀澄清。
1. 首先保证母液是澄清的;
2.
一定要按照顺序依次将溶剂加入,进行下一步操作之前必须保证上一步操作得到的是澄清的溶液,可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
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