| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 577/602 |
| MW | 686.25 |
| CAS# | N/A |
| Solvent | DMSO |
| Storage | F/D/L |
| Category | CellBIOLOGy LabelingCells |
| Related | ViABIlityandProliferation CellViability MDRResearch BiochemicalAssays |
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.
| References&Citations |  PrinterFriendlyVersion |
- ZhangC,JinS,YangK,XueX,LiZ,JiangY,ChenWQ,DaiL,ZouG,LiangXJ.(2014)Cellmembranetrackerbasedonrestrictionofintramolecularrotation.ACSApplMaterInterfaces,6,8971.
- RezatofighiSH,GouldS,VoBN,MeleK,HughesWE,HartleyR.(2013)Amultiplemodelprobabilityhypothesisdensitytrackerfortime-lapsecellmicroscopysequences.InfProcessMedImaging,23,110.
- KleinJ,LeupoldS,BieglerI,BiedendieckR,MunchR,JahnD.(2012)TLM-Tracker:softwareforcellsegmentation,trackingandlineageanalysisintime-lapsemicroscopymovies.Bioinformatics,28,2276.
- FangCY,VaijayanthimalaV,ChengCA,YehSH,ChangCF,LiCL,ChangHC.(2011)Theexocytosisoffluorescentnanodiamondanditsuseasalong-termcelltracker.Small,7,3363.
- ChalfounJ,CardoneA,DimaAA,AllenDP,HalterMW.(2010)Overlap-BasedCellTracker.JResNatlInstStandTechnol,115,477.
- SwirskiFK,BergerCR,FigueiredoJL,MempelTR,vonAndrianUH,PittetMJ,WeisslederR.(2007)Anear-infraredcelltrackerreagentformultiscopicinvivoimagingandquantificationofleukocyteimmuneresponses.PLoSOne,2,e1075.
- Ionescu-ZanettiC,WangLP,DiCarloD,HungP,DiBlasA,HugheyR,LeeLP.(2005)Alkalinehemolysisfragilityisdependentoncellshape:resultsfromamorphologytracker.CytometryA,65,116.
- Caetano-PintoP,JanssenMJ,GijzenL,VerscheijdenL,WilmerMJ,MasereeuwR.(2016)Fluorescence-BasedTransportAssaysRevisitedinaHumanRenalProximalTubuleCellLine.MolPharm,13,933.
- MannaA,DeSarkarS,DeS,BauriAK,ChattopadhyayS,ChatterjeeM.(2015)ThevariablechemotherapeuticresponseofMalabaricone-Ainleukemicandsolidtumorcelllinesdependsonthedegreeofredoximbalance.Phytomedicine,22,713.
- BeemE,SegalMS.(2013)Evaluationofstabilityandsensitivityofcellfluorescentlabelswhenusedforcellmigration.JFluoresc,23,975.
