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主营:研究并生产荧光和发光探针,信号转导研究的试剂
℡ 4000-520-616
℡ 4000-520-616
AAT Bioquest/Cell Meter™ 2-NBDG Glucose Uptake Assay Kit/23500/200 Tests
产品编号:23500
市  场 价:¥143560.00
场      地:美国(厂家直采)
联系QQ:1570468124
电话号码:4000-520-616
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美  元  价:$7178.00
品      牌: AAT Bioquest
公      司:AAT Bioquest
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AAT Bioquest/Cell Meter™ 2-NBDG Glucose Uptake Assay Kit/23500/200 Tests
商品介绍
Overview
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Ex/Em(nm)485/540
MWN/A
CAS#186689-07-6
SolventDMSO
StorageF/D/L
CategoryTransporters
GlucoseTransporters
RelatedCellFunctionalAnalysis
Glucosemetabolism,aprocesswhichconvertsglucoseintoenergy,isaprimarysourceofenergysupplyinmostorganisms.2-NBDG[2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose],afluorescentlytaggedglucosetracer,hasbeenproventoeffectivelymonitorglucosetransportationincells,as2-NBDGtransportsintocellsbythesameglucosetransporters(GLUTs)asglucose.Once2-NBDGisuptakenincells,itundergoesphosphorylationatC-6positiontogive2-NBDG-6-phosphate,whichiswellretainedwithinthecells.Comparedtootherglucosetracers,suchas2-DGorFDG,2-NBDGallowsinsitumeasurementsof2-NBDGwithhightemporalandspatialresolutionatsinglecelllevel.AATBioquest"sCellMeter™2-NBDGGlucoseUptakeAssayKitprovidesasensitiveandnon-rADIoactiveassayformeasuringglucoseuptakeinculturedcells.Inthiskit,AssayBufferIisusedtoenhancetheuptakeandretentionof2-NBDGincells,whileAssayBufferIIcanimprovethesignal-to-backgroundratioof2-NBDGinthecells.ThefluorescencesignalcanbemonitoredbyfluorescencemicroscopeorflowcytometerwithaFITCfilterset.CellMeter™2-NBDGGlucoseUptakeAssayKitisthemostrobusttoolformonitoringglucosetransporters.
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Thisprotocolonlyprovidesaguideline,andshouldbemodifiedaccordingtoyourspecificneeds.
  1. Preparecells:
    1. Foradherentcells:Platecellsovernightingrowthmediumat10,000to40,000cells/well/100μLfora96-wellplateor2,500to10,000cells/well/20μLfora384-wellplatewithblackwallandclearbottom.
    2. Fornon-adherentcells:CentrifugethecellsfromtheculturemediumandsUSPendthecellpelletsinculturemediumat50,000-100,000cells/well/100µLfora96-wellpoly-Dlysineplateor10,000-25,000cells/well/20µLfora384-wellpoly-Dlysineplate.
    3. Addtestcompoundsintothecellsandincubateforadesiredperiodoftime(suchas24,48or96hours)ina37°C,5%CO2incubator.Forblankwells(mediumwithoutthecells),addthesameamountofcompoundbuffer.
      Note:Eachcelllineshouldbeevaluatedonanindividualbasistodeterminetheoptimalcelldensityandincubationtime.WeincubatedCHO-K1cellswith20mMGlucoseforglucosecompetitionassay,and100µMPhloretinforGLUTsinhibitionassay.SeeFigure2fordetails.

  2. Staincells:
    1. Preparestainingsolutionbyadding5μLof2-NBDG(10mg/mL)(ComponentA)to1.5mLofAssayBufferI(ComponentB).Thedye-loadingsolutionisstablefor1houratroomtemperature,protectedfromlight.
      Note:Astheoptimalstainingconditionsmayvarydependingondifferentcelltypes,it’srecommendedtodeterminetheoptimalconcentrationofComponentAforeachspecificexperiment.
    2. Attheendofthetreatment,centrifugetheplatefor5minutesat800rpmwithbrakeoffpriortoyourexperiment.Aspiratethesupernatantwithoutdisturbingcells.
    3. Add100µL/well(96-wellplate)or25µL/well(384-wellplate)ofstainingsolution(fromStep2.1).
      Note:Opticalincubationtimewillneedtobedeterminedforeachcelllineandforeachspecificexperiment.WeincubatedCHO-K1cellswith100µM2-NBDG(~34µg/mL)for20minutestoshowsufficientglucoseuptake.Seefigure2fordetails.
    4. Attheendoftheincubation,centrifugetheplatefor5minutesat800rpm.Removestainingsolutionwithoutdisturbingcells.
    5. Forfluorescencemicroscope:WashcellswithAssayBufferI(ComponentB)once.Keepcellsin100µL/well(96-wellplate)or25µL/well(384-wellplate)ofAssayBufferII(ComponentC).MonitorthefluorescencesignalusingafluorescencemicroscopewithFITCfilter.
    6. Forflowcytometer:DetachcellsifrequiredusingEDTAandresuspendcellsin100µL/sampleofAssayBufferI(ComponentB).MonitorthefluorescencesignalusingaflowcytometeratFITCchannel.
References&Citations
CitationExplorer

NT1014,anovelbiguanide,inhibitsovariancancergrowthinvitroandinvivo
Authors:LuZhang,JianjunHan,AmandaLJackson,LeslieNClark,JoshuaKilgore,HuiGuo,NickLivingston,KennethBatchelor,YajieYin,TimothyPGilliam
Journal:JournalofHematology&Oncology(2016):91

GlutaminepromotesovariancancercellproliferationthroughthemTOR/S6pathway
Authors:LingqinYuan,XiuguiSheng,AdamKWillson,DarioRRoque,JessicaEStine,HuiGuo,HannahMJones,ChunxiaoZhou,VictoriaLBae-Jump
Journal:Endocrine-relatedcancer(2015):577--591


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