2nbdg用于乳腺癌mdamb231细胞荧光检测研究

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1、2NBDG用于乳腺癌MDAMB231细胞荧光检测研究作者：胡慧单秀红朱伟钱晖许文荣王亚非【摘要】　　目的：评价荧光2N［7硝基苯2乙二酸，34羟氨基］(2［N(7nitrobenz2oxa1，3diazol4yl)amino］，NBD)标记2脱氧葡萄糖（2deoxyglucose，2DG）即2NBDG，靶向探测高表达葡萄糖转运蛋白1（Glut1）的MDAMB231乳腺癌细胞可行性。方法：2NBDG分别孵育接种在6孔板里的人乳腺癌MDAMB231细胞1,10,20,40min后，荧光显微镜观察并摄像；为了进行竞争性抑制,用D型葡萄糖预先孵育

2、细胞30min后再加入2NBDG孵育20min，荧光显微镜观察并摄像。同时用流式细胞仪分析孵育不同时间及有无竞争抑制的细胞吸收2NBDG量的差异。结果：荧光成像显示2NBDG积聚在MDAMB231细胞膜和细胞质内，加入D型葡萄糖竞争抑制后，荧光信号显著减弱；流式细胞仪分析表明最初的1min2NBDG被肿瘤细胞迅速摄取（MDAMB231细胞自发荧光强度为2.66±0.09，2NBDG孵育1min后荧光强度为4.04±0.18），10min时吸收持续增加（荧光强度为5.64±0.17），20min吸收最明显（荧光强度为25.10±0.57）；加入D型葡萄糖竞争抑制后，细胞摄取

3、2NBDG的荧光强度降低46%。结论:2NBDG能迅速被高表达Glut1的MDAMB231乳腺癌细胞靶向吸收，有望成为葡萄糖代谢的光学标记物来检测高代谢的恶性肿瘤。12【关键词】乳腺癌细胞；2NBDG；荧光成像；流式细胞仪　　［Abstract］Objective:Toassessthefeasibilityof2deoxyglucoselabeledwiththefluorescence(2［N(7nitrobenz2oxa1，3diazol4yl)amino］,NBD)whichis2NBDG,asatargetingprobefordetectingb

4、reastcancercellswhichhighlyexpressesGlucosetransporter1(Glut1).Methods：MDAMB231cellswhichweregrownin6wellplatesincubatedby2NBDGfor1min,10min,20minand40minrespectively,andtheresultwereanalyzedbyfluorescencemicroscopyimaging.Forcompetitiveinhibition,thecellswerepreincubatedbyDglucosefor30minut

5、esbeforeadding2NBDGincubatedfor20minutes,thentheresultwereanalyzedbyfluorescencemicroscopyimaging.Also,theamountdifferencesofabsorptionof2NBDGamongcellsincubatedfordifferenttimeandwithorwithoutcompetitiveinhibitioncomparedbyflowcytometry.Results:Fluorescencemicroscopyimagingdisplayed2NBDGaccumu

6、latedinMDAMB231cellsmembraneandcytoplasm,andthefluorescentsignalmarkedlydroppedasaddingfreeDglucoseforcompetitiveinhibition.Flowcytometryanalysisof2NBDGuptakedisplayedrapiduptakeforthefirstoneminute(the12FluorescencebackgroundofMDAMB231cellswas2.66±0.09,incubationwith2NBDGfor1minutewas4.04±0

7、.18),theabsorptionof2NBDGincubationfor10minutescontinuedtoincreasely(fluorescenceintensitywas5.64±0.17),themostobviousabsorptionwasincubationfor20minutes(fluorescenceintensityof25.10±0.57),andadditionofDglucoser