2-NBDG,葡萄糖攝取熒光探針
2-NBDG,葡萄糖攝取熒光探針(2-NBD-Glucose)是熒光標(biāo)記的脫氧葡萄糖類似物,用于直接監(jiān)測(cè)活細(xì)胞和組織對(duì)葡萄糖的攝取。 2-NBD用作檢測(cè)瘤形成的局部造影劑。 攝取可以通過(guò)實(shí)時(shí)共聚焦、高分辨率或?qū)捯晥?chǎng)熒光顯微鏡和流式細(xì)胞術(shù)進(jìn)行監(jiān)測(cè)。
溶解度Solubility:溶于乙醇至 50 mM,溶于 DMSO 至 25 mM
性狀:solid,固體粉末
儲(chǔ)藏條件:4℃ (1-2weeks), longer storage period at -20℃ (1-2years)
1. 2-NBDG儲(chǔ)存液制備方法?
向 1mg粉末中加 100μl 無(wú)水DMSO 或者 乙醇,進(jìn)行充分溶解混勻,制備成10mg/ml的儲(chǔ)存液,按照單次用量進(jìn)行分裝后,在-20C避光條件下冷藏存儲(chǔ)。若已經(jīng)配制成水溶液后,建議存周期不超過(guò)一天。
如實(shí)驗(yàn)需要延長(zhǎng)保存周期,最好是以粉末的形式在-20C條件下保存。先用乙醇溶解固體粉末,然后分裝成小量裝入獨(dú)立瓶?jī)?nèi),使用真空泵將溶劑抽干即可。此時(shí)不推薦用DMSO或DMF為溶劑,因揮發(fā)性比較差。
2. 2-NBDG工作濃度?
2-NBDG 的常用工作濃度大約10-600μM。細(xì)菌或者酵母菌染色的建議濃度范為100-600μM,原代和傳代培養(yǎng)哺乳動(dòng)物細(xì)胞建議工作濃度范圍為10-200μM。
日常實(shí)驗(yàn)過(guò)程 2-NBDG 工作濃度范圍為: 30-300μM。
細(xì)胞需要在在合適的溫度條件下孵育,以充分檢測(cè)綠-黃色熒光。攝取能力強(qiáng)弱受溫度影響,孵育時(shí)間有幾秒鐘,到30min,甚至更長(zhǎng),取決于探針終濃度、細(xì)胞類型、培養(yǎng)條件和一些其他因素。
2-NBDG案例:
Assessment of 2-NBDG as a marker of glucose metabolism in animals. a Multispectral fluorescence imaging of HT-29 subcutaneous tumors in nude mice 40 min following administration of 500 nmol of 2-NBDG in fed versus fasting states. Red arrows indicate the location of the tumor. There is a noticeable increase in 2-NBDG signal after the animal was kept NPO for 24 h. b Analysis of n =4 animals with subcutaneous tumors imaged, while fed and fasting shows a statistically significant p 0.05 increase in 2-NBDG uptake in fasting animals. c Statistically significant p 0.05 increase in 2-NBDG signal in the brain of an animal experiencing generalized seizures induced by injection of pilocarpine i.p. versus control. These data suggest that 2-NBDG localization reflects true glucose utilization rather than nonspecific extravasation.??