Overview | PrinterFriendlyVersion |
Ex/Em(nm) | 540/590 |
MW | N/A |
CAS# | N/A |
Solvent | N/A |
Storage | F/D/L |
Category | CellBIOLOGy CellMetabolism |
Related | RedoxEnzymes |
Spectrum | AdvancedSpectrumViewer |
- Preparecells:
- Foradherentcells:Platecellsovernightingrowthmediumat10,000to40,000cells/well/90μLfora96-wellplateor2,500to10,000cells/well/20μLfora384-wellplate.
- Fornon-adherentcells:CentrifugethecellsfromtheculturemediumandsUSPendthecellpelletsinculturemediumat100,000-200,000cells/well/90µLfora96-wellpoly-Dlysineplateor25,000-50,000cells/well/20µLfora384-wellpoly-Dlysineplate.Centrifugetheplateat800rpmfor2minuteswithbrakeoffpriortoyourexperiment.
Note:Eachcelllineshouldbeevaluatedonanindividualbasistodeterminetheoptimalcelldensity.
- Prepareworkingsolution:
- Thawallthekitcomponentatroomtemperaturebeforeuse.
- MakeJZL1707NAD(P)HSensorworkingsolution:Add10µLofJZL1707NAD(P)Hsensorstocksolution(ComponentA)into2.5mLofAssayBuffer(ComponentB),andmixwell.Theworkingsolutionisstablewithin1houratroomtemperature.
Note:40µLofJZL1707NAD(P)Hsensorstocksolutionisenoughforoneplate.AliquotedandstoredunusedJZL1707NAD(P)Hsensorstocksolutionat≤-20ºC.Protectitfromlightandavoidrepeatedfreeze-thawcycles.
- RuntheNADH/NADPHassay:
- TostimulateNADP/NADPH,treatcellswith10μLof10Xtestcompounds(96-wellplate)or5μLof5Xtestcompounds(384-wellplate)inserumfreemediumoryourdesiredbuffer(suchasPBSorHHBS).Forcontrolwells(untreatedcells),addthecorrespondingamountofmediumorcompoundbuffer.
Note:JZL1707NAD(P)Hsensorisserumsensitive,thereforeit’srecommendedtokeepcellsinserum-freemediumorthebufferofyourchoice.Alternatively,cellscanbepreparedandtreatedinregularfullmedium.ChangetoserumfreemediumorbufferofyourchoicewhenincubationwithJZL1707NAD(P)Hsensor. - Add100µL/well(96-wellplate)or25µL/well(384-wellplate)ofJZL1707NAD(P)Hsensorworkingsolution(fromStep2.2)inthecellplate.Co-incubatecellswithtestcompoundandJZL1707NAD(P)Hsensorworkingsolutionat37ºCfor30-60minutes,protectedfromlight.
Note:ForaNADH/NADPHpositivecontroltreatment:HeLacellswereincubatedwith100µMNADHorNADPHfor30minutesinserum-freemedium,andco-incubatedwithJZL1707NAD(P)Hsensorworkingsolutionat37ºCforanother30minutes.SeeFigure1fordetails. - Washcellswithyourdesiredbufferonce.RemovesolutionineachwellandaddAssayBuffer(ComponentB)100µL/wellfora96-wellplateor25µL/wellfora384-wellplate.
- MonitorthefluorescenceincreaseusingmicroplatereaderatEx/Em=540/590nm(cutoff=570nm)withbottomreadmode,ortakeimagesusingfluorescencemicroscopewithaCy3®filter.
- TostimulateNADP/NADPH,treatcellswith10μLof10Xtestcompounds(96-wellplate)or5μLof5Xtestcompounds(384-wellplate)inserumfreemediumoryourdesiredbuffer(suchasPBSorHHBS).Forcontrolwells(untreatedcells),addthecorrespondingamountofmediumorcompoundbuffer.
References&Citations | PrinterFriendlyVersion |
1. RubinTan,JianshaLi,XiaochunPeng,LipingZhu,LeiCai,TaoWang,YuanSu,KaikobadIrani,andQinghuaHu.GAPDHiscriticalforsuperiorefficacyoffemalebonemarrow-derivedmesenchymalstemcellsonpulmonaryhypertension.CardiovascRes,Oct2013;100:19-27.
2. StephenY.Xue,ValeriaY.Hebert,DaniciaM.Hayes,CorieN.Robinson,MitziGlover,andTammyR.Dugas.NucleosideReverseTranscriptaseInhibitorsInduceaMitophagy-AssociatedEndothelialCytotoxicityThatIsReversedbyCoenzymeQ10Cotreatment.Toxicol.Sci.,Aug2013;134:323–334.
3. KateJ.Roberts,AndrewCross,OlgaVasieva,RobertJ.Moots,andStevenW.Edwards.Inhibitionofpre-Bcellcolony-enhancingfactor(PBEF/NAMPT/visfatin)decreasestheABIlityofhumanneutrophilstogeneratereactiveoxidantsbutdoesnotimpairbacterialkilling.J.Leukoc.Biol.,Sep2013;94:481-492.
4. WeijingCai,MayaRamdas,LiZhu,XueChen,GaryE.Striker,andHelenVlassara.Oraladvancedglycationendproducts(AGEs)promoteinsulinresistanceanddiabetesbydepletingtheantioxidantdefensesAGEreceptor-1andsirtuin1.PNAS,Sep2012;109:15888-15893.
5. YueQiu,ClausTittiger,ClaudeWicker-Thomas,GaëlleLeGoff,SharonYoung,EricWajnberg,ThierryFricaux,NathalieTaquet,GaryJ.Blomquist,andRenéFeyereisen.Aninsect-specificP450oxidativedecarbonylaseforcuticularhydrocarbonbiosynthesis.PNAS,Sep2012;109:14858-14863.
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颜料是用着色的物质通常与被着色的物质混合在一起主要以无机化合物为主;染料是一种用来直接或经媒染剂作用而能附着在各种纤维和其他材料上的有色物质有的可以跟被染物质化合,多以有机化合物为主。 颜料不能上色,而染料能上色. 颜料和染料的区别 颜料是一种微细粉末状的有色物质,一般不溶于水、油和溶剂,但能均匀的分散在其中。颜料是色漆的次要成膜物质,在木材装饰过程中调制底漆、腻子以及木才着色,也经常使用颜料.
想请问一下,DAPI这个染料到底有没有膜通透性,我通过百度搜索查询关于DAPI染料的,基本上是说它能透过细胞膜对活细胞和死细胞均能染上蓝色;但是也有人说DAPI只可以透过死细胞膜,不能对活细胞进行染色,用以区分活死细胞,到底哪个是对的啊,蒙了!!!!!!
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是欢快活泼的光辉色彩,是暖色系中最温暖的色,它使人联想到金色的秋天,丰硕的果实;
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