![AAT Bioquest/FDA[二醋酸荧光素]*CAS 596-09-8*/22020/1 g](https://www.ebiomall.cn/images/AAT201712/chemical-structure-for-fda-fluorescein-diacetate-cas-596-09-8.png)
| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 494/521 |
| MW | 416.38 |
| CAS# | 596-09-8 |
| Solvent | DMSO |
| Storage | F/D/L |
| Category | CellBIOLOGy LabelingCells |
| Related | ViABIlityandProliferation CellViability BiochemicalAssays |
| Spectrum | AdvancedSpectrumViewer |
1.Prepare2-10mMDMSOstocksolution
For#22014add45uLDMSOintoa50µgvialtomake2 mMstocksolution(1mg/mlisequivalentto1.8mM);
For#22015add36uLDMSOintoa50µgvialtomake10mMstocksolution(1mg/mlisequivalentto1.46mM);
For#22016add153uLmlDMSOtomake10mMstocksolution(1mg/mlisequivalentto1.53mM);
For#22017add215uLmlDMSOtomake10mMstocksolution(1mg/mlisequivalentto2.15mM);
For#22020dissolve4.2mgin1mlDMSOtomake10mMstocksolution(1mg/mlisequivalentto~2.4mM);
Note:Thestocksolutionshouldbeusedpromptly;anyremainingsolutionshouldbealiquotedandfrozenat<-20oC.Avoidrepeatedfreeze-thawcycles,andprotectfromlight.
2.Preparedyeworkingsolution
Preparea1to20µMdyeworkingsolutionrightbeforeusebydilutingtheDMSOstocksolutionfromStep1withHanksand20mMHepesbuffer(HHBS)orthebufferofyourchoice,pH7.Mixthemwellbyvortexing.
3.Analyzecellswithaflowcytometerorafluorescencemicroscope:
3.1 Treatcellswithtestcompoundsforadesiredperiodoftime.
3.2 Centrifugethecellstoget2-10x105cellspertube.
3.3 ResUSPendcellsin500µLofthedyeworkingsolution(fromStep2).
3.4 Incubatecellswithadyesolutionatroomtemperatureor37°Cfor15to30min,protectedfromlight.
3.5 Removethedyeworkingsolutionfromthecells,washthecellswithHHBSorbufferofyourchoice.Resuspendcellsin500µLofpre-warmedHHBSormediumtoget2-10x105cellspertube.
3.6 MonitorthefluorescencechangeatEx/Em=490/520nmwithaflowcytometer(FL1channel)orafluorescencemicroscope.
Note:Forbacterialcellsstaining:Stainingismostefficientwhenstocksolutionisdiluted1:800innutrientbrothpreconditionedbyovernightgrowthofthetestbacteria,butfreshnutrientbrothorPBSmayalsobeused.BacterialsuspensionsshouldbedilutedwithPBSto105-107organismsperml.Bacteriamaybestainedbyapplyingonemlofsolutionto.45µmfilter(25mm)andvacuumfilteringtoremovesolution,thenadding1mlofdyesolutionandincubating5-10minutesatroomtemperature.
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