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AAT Bioquest/Cell Explorer™ Live Cell Tracking Kit *Orange Fluorescence*/22622/200 Tests
| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 528/541 |
| MW | N/A |
| CAS# | N/A |
| Solvent | N/A |
| Storage | F/D/L |
| Category | CellBIOLOGy LabelingCells |
| Related | FluorescenceImaging BiochemicalAssays |
| Spectrum | AdvancedSpectrumViewer |
1.PrepareCells:
1.1 Foradherentcells:Platecellsovernightingrowthmediumat10,000to40,000cells/well/90µLfor96-wellplatesor2,500to10,000cells/well/20µLfor384-wellplates.
1.2 Fornon-adherentcells:CentrifugethecellsfromtheculturemediumandthensUSPendthecellpelletsinculturemediumat50,000-100,000cells/well/90µLfor96-wellpoly-Dlysineplatesor10,000-25,000cells/well/20µLfor384-wellpoly-Dlysineplates.Centrifugetheplatesat800rpmfor2minuteswithbrakeoffpriortotheexperiment.
Note:Eachcelllineshouldbeevaluatedonanindividualbasistodeterminetheoptimalcelldensity.
2.Prepare10XTrackIt™Orangestainsolution:
Dilute500XTrackIt™OrangeDMSOstocksolution(ComponentA)intoAssayBuffer(ComponentB)tomakea10to25XTrackIt™Orangeworkingsolution.Theworkingsolutionshouldbepreparedenoughforallthewellsat10μL/wellwiththeappropriateconcentration.Forexample,togeta1XfinalconcentrationofTrackIt™Orangeforone96-wellmicroplate,dilute20μLoftheTrackIt™OrangeDMSOstocksolutioninto1mLofAssayBuffer(ComponentB)tomake1mLof10XTrackIt™Orangeworkingsolution.
Note1:TheunusedportionoftheTrackIt™Orangestocksolutionshouldbestoredat-20oC.Avoidrepeatedfreeze/thawcycles.
Note2:ThefinalconcentrationoftheTrackIt™Orangeworkingsolutionshouldbeempiricallydeterminedfordifferentcelltypesand/orexperimentalconditions.Itisrecommendedtotestattheconcentrationsthatareatleastoveratenfoldrange.
3.Stainthecells:
3.1 Tothecellwellsadd10XTrackIt™orangeworkingsolution(fromStep2)whichshouldbeequalto1/10ofthevolumeofcellculturemedium.Forexample,fora96-wellplate,add10µL/wellof10XTrackIt™Orangeworkingsolutionintothecells.
3.2 Incubatethecellsina37oC,5%CO2incubatorfor15minutesto1hour.
3.3 WashcellswithHanksand20mMHepesbuffer(HBSS)oranappropriatebuffer.
3.4 Fillthecellwellswithgrowthmedium.
3.5 AnalyzethecellsusingafluorescencemicroscopeorflowcytometerwithTRITCfiltersets(Ex/Em=540/570nm).
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