| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 646/659 |
| MW | N/A |
| CAS# | N/A |
| Solvent | N/A |
| Storage | F/D/L |
| Category | CellAnalysis CellCytotoxicity |
| Related | ViABIlityandProliferation CellViability BiochemicalAssays |
| Spectrum | AdvancedSpectrumViewer |
1.Preparecells:
Plate100to100,000×10cellsperwellinatissueculturemicroplatewithblackwallandclearbottom.Addtestcompoundsintothecellsandincubateforadesiredperiodoftime(suchas24,48or96hours)ina37°C,5%CO2incubator.Forblankwells(mediumwithoutthecells),addthesameamountofcompoundbuffer.Thesuggestedtotalvolumeis100µL/well/96-wellplate,and25µL/well/384-wellplate.
Note:Eachcelllineshouldbeevaluatedontheindividualbasistodeterminetheoptimalcelldensityforproliferationorcytotoxicityinduction.Forproliferationassays,usefewercells;forcytotoxicityassays,usemorecellstostartwith.
2.Preparedye-loadingsolution:
2.1 Thawoneofeachkitcomponentatroomtemperaturebeforeuse.
2.2 MakeCytoCalcein™NIRstocksolution:Add20µLofDMSO(ComponentB)intothevialofCytoCalcein™NIR(ComponentA),andmixthemwell.
Note:20µLofCytoCalcein™NIRstocksolutionisenoughforoneplate.UnusedCytoCalcein™NIRstocksolutioncanbealiquotedandstoredat<-20oCforonemonthifthetubesaresealedtightly.Protectfromlightandavoidrepeatedfreeze-thawcycles.
2.3 MakeCytoCalcein™NIRdye-loadingsolutionforonecellplate:Add20µLofCytoCalcein™NIRstocksolution(fromStep2.2)intothebottleofAssayBuffer(10mL,ComponentC),andmixthemwell.Thedye-loadingsolutionisstableforatleast2hoursatroomtemperature.
3.Runthecellviabilityassay:
3.1 Treatcellswithtestcompoundsasdesired(fromStep1).
Note:Itisnotnecessarytowashcellsbeforeaddingcompound.However,iftestedcompoundsareserumsensitive,growthmediumandserumfactorscanbeaspiratedawaybeforeaddingcompounds.Add100µL/well(96-wellplate)and25µL/well(384-wellplate)of1XHank’ssaltsolutionand20mMHepesbuffer(HHBS)orthebufferofyourchoiceafteraspiration.Alternatively,cellscanbegrowninserum-freemedia.
3.2 Removethemediumfromthecells.
Note:Mediummustberemovedbeforedyeloading.
3.3 Add100µL/well(96-wellplate)or25µL/well(384-wellplate)ofdye-loadingsolution(fromStep2.3).
3.4 Incubatethedye-loadingplateatroomtemperatureor37oCfor1hour,protectedfromlight.
Note1:Theappropriateincubationtimedependsontheindividualcelltypeandcellconcentrationused.Optimizetheincubationtimeforeachexperiment.
Note2:Fornon-adherentcells,itisrecommendedtocentrifugecellplatesat800rpmfor2minuteswithbrakeoffafterincubation.
3.5 MonitorthefluorescenceintensityatEx/Em=635/670nm.
| References&Citations |  CitationExplorer |
Functionalevidencethattheself-renewalgeneNANOGregulatesesophagealsquamouscancerdevelopment
Authors:DengLi,XiaocongXiang,FeiYang,DongqinXiao,KangLiu,ZhuChen,RuolanZhang,GangFeng
Journal:BiochemicalandBiophysicalResearchCommunications(2017)
NINJ2--Anovelregulatorofendothelialinflammationandactivation
Authors:JingjingWang,JingjingFa,PengyunWang,XinzhenJia,HuixinPeng,JingChen,YifanWang,ChenhuiWang,QiuyunChen,XinTu
Journal:CellularSignalling(2017)
ErythropoietinStimulatesEndothelialProgenitorCellstoInduceEndothelializationinanAneurysmNeckAfterCoilEmbolizationbyModulatingVascularEndothelialGrowthFactor
Authors:PeixiLiu,YingjieZhou,QingzhuAn,YayingSong,XiChen,Guo-YuanYang,WeiZhu
Journal:MEDICINE(2016):1--8
FlexIBLeEndoscopicSprayApplicationofRespiratoryEpithelialCellsasPlatformTechnologytoApplyCellsinTubularOrgans
Authors:AnjaLenaThiebes,ManuelArminReddemann,JohannesPalmer,ReinholdKneer,StefanJockenhoevel,ChristianGabrielCornelissen
Journal:TissueEngineeringPartC:Methods(2016):322--331
Influenceofhypothermiaandsubsequentrewarminguponleukocyte-endothelialinteractionsandexpressionofJunctional-Adhesion-MoleculesAandB
Authors:NicolaiVBogert,IsabellaWerner,AngelaKornberger,PatrickMeybohm,AntonMoritz,TillKeller,UlrichAStock,AndresBeiras-Fernandez
Journal:Scientificreports(2016)
Influenceofhypothermiaandsubsequentrewarminguponleukocyte-endothelialinteractionsandexpressionofJunctional-Adhesion-MoleculesAandB
Authors:NicolaiVBogert,IsabellaWerner,AngelaKornberger,PatrickMeybohm,AntonMoritz,TillKeller,UlrichAStock,AndresBeiras-Fernandez
Journal:Scientificreports(2016)
InhibitionofABCtransportproteinsbyoilsandsprocessaffectedwater
Authors:HattanAAlharbi,DavidMVSaunders,AhmedAl-Mousa,JaneAlcorn,AlbertoSPereira,JonathanWMartin,JohnPGiesy,SteveBWiseman
Journal:AquaticToxicology(2016):81--88
Rapidgenerationofcollagen-basedmicrotissuestostudycell--matrixinteractions
Authors:Marie-ElenaBrett,AlexandraLCrampton,DavidKWood
Journal:Technology(2016):1--8
ToxicokineticsandtoxicodynamicsofchlorpyrifosisalteredinembryosofJapanesemedakaexposedtooilsandsprocess-affectedwater:evidenceforinhibitionofP-glycoprotein
Authors:HattanAAlharbi,JaneAlcorn,AhmedAl-Mousa,JohnPGiesy,SteveBWiseman
Journal:JournalofAppliedToxicology(2016)
Sprayingrespiratoryepithelialcellstocoattissue-engineeredconstructs
Authors:AnjaLenaThiebes,StefanieAlbers,ChristianKlopsch,StefanJockenhoevel,ChristianGCornelissen
Journal:BioResearchopenaccess(2015):278--287
