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主营:研究并生产荧光和发光探针,信号转导研究的试剂
℡ 4000-520-616
℡ 4000-520-616
AAT Bioquest/DiSBAC2(3) [Bis-(1,3-diethylthiobarbituric acid)trimethine oxonol]/21414/25 mg
产品编号:21414
市  场 价:¥27560.00
场      地:美国(厂家直采)
联系QQ:1570468124
电话号码:4000-520-616
邮      箱: info@ebiomall.com
美  元  价:$1378.00
品      牌: AAT Bioquest
公      司:AAT Bioquest
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AAT Bioquest/DiSBAC2(3) [Bis-(1,3-diethylthiobarbituric acid)trimethine oxonol]/21414/25 mg
商品介绍
Overview
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Ex/Em(nm)535/560
MW436.55
CAS#N/A
SolventDMSO
StorageF/D/L
CategoryIonChannels
MembranePotentials
RelatedNeurologicalStains
BiochemicalAssays
DiSBAC2(3)isasensitiveslow-responsemembranepotentialprobethatiswidelyusedformeasuringmembranepotentialsofmanyBIOLOGicalsystems.Ingeneral,slow-responseprobesexhibitpotential-dependentchangesintheirtransmembranedistributionthatareaccompaniedbyafluorescencechange.Themagnitudeoftheiropticalresponsesismuchlargerthanthatoffast-responseprobes(typicallya1%fluorescencechangepermV).Slow-responseprobes,whichincludecationiccarbocyanines,rhodaminesandanionicoxonols,aresuitablefordetectingchangesinaveragemembranepotentialsofnonexcitablecellscausedbyrespiratoryactivity,ion-channelpermeABIlity,drugbindingandotherfactors.
SpectrumAdvancedSpectrumViewer

Sorry,yourbrowserdoesnotsupportinlineSVG.RelativeIntensity(%)100806040200Sorry,yourbrowserdoesnotsupportinlineSVG.
Sorry,yourbrowserdoesnotsupportinlineSVG.Sorry,yourbrowserdoesnotsupportinlineSVG.
Movemouseovergridtodisplaywavelength&intensityvalues.

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Thisprotocolonlyprovidesaguideline,andshouldbemodifiedaccordingtoyourspecificneeds.

1.Preparecells:

1.1   Foradherentcells:Platecellsovernightingrowthmediumat40,000to80,000cells/well/100µLfor96-wellplatesor10,000to20,000cells/well/25µLfor384-wellplates.

 

1.2   Fornon-adherentcells:CentrifugethecellsfromtheculturemediumandthensUSPendthecellpelletsinequalamountofHHBSandMPdye-loADIngsolution(seeStep2.2below)at125,000to250,000cells/well/100µLfor96-wellpoly-Dlysineplatesor30,000to60,000cells/well/25µLfor384-wellpoly-Dlysineplates.Centrifugetheplatesat800rpmfor2minuteswithbrakeoffbeforetheexperiments.

Note:Eachcelllineshouldbeevaluatedonanindividualbasistodeterminetheoptimalcelldensityfortheintracellularcalciummobilization.

 

2.PrepareDiSBAC2(3)dye-loadingsolution(for1plate):

2.1   Preparea10to30mMstocksolutionofDiSBAC2(3)inhigh-quality,anhydrousDMSO.Thestocksolutionshouldbeusedpromptly;anyremainingsolutionneedbealiquotedandfrozenat<-20oC.

Note:Avoidrepeatedfreeze-thawcycles,andprotectfromlight.

2.2   Preparea2XDiSBAC2(3)dye-loadingsolution:Onthedayoftheexperiment,eitherdissolveDiSBAC2(3)solidinDMSOorthawanaliquotoftheDiSBAC2(3)stocksolutiontoroomtemperature.Preparea2Xworkingsolutionof20to40µMinHanksand20mMHepesbuffer(HHBS)orbufferofyourchoice,pH7with0.04%to0.08%Pluronic®F-127(Cat.#20053)and2mMTrypanRedPlus™(Cat.#2456).Mixthemwellbyvotexing.Thisworkingsolutionisstableforatleast2hoursatroomtemperature.

 

3.RunMembranePotentialAssay:

3.1   Add100µL/well(96-wellplate)or25µL/well(384-wellplate)DiSBAC2(3)dye-loadingsolution(fromStep2.2)intothecellplate.

Note1:Ifyourscreencompoundsinterferewithgrowthmediumandserumfactors,replacethegrowthmediumwithequalvolumeofHHBSbufferbeforeaddingtheDiSBAC2(3)dye-loadingsolution.Alternatively,cellscanbegrowninserum-freeconditions.

Note2:DoNOTwashthecellsafterdyeloading.

 

3.2   Incubatethedye-loadingplateinacellincubatorfor30to60minutes.

Note:Insomecases,incubationatroomtemperaturefor30to60minmayworkbetter.

 

3.3   PreparethecompoundplatesbyusingHHBSoryourdesiredbuffer.

 

3.4   RunthemembranepotentialassaybymonitoringthefluorescenceintensityatEx/Em=540/590nm(Cat.# 21414).

Note:Itisimportanttorunthesignaltestbeforeyourexperiment.Differentinstrumentshavetheirownintensityrange.Adjustthesignaltestintensitytothelevelof10%to15%ofthemaximuminstrumentintensitycounts.Forexample,themaximumfluorescenceintensitycountforFLIPR-384is65,000,sotheinstrumentsettingsshouldbeadjustedtohaveitssignaltestintensityaround7,000to10,000.

References&Citations
CitationExplorer

SuppressionofKV7/KCNQpotassiumchannelenhancesneuronaldifferentiationofPC12cells
Authors:NajingZhou,ShaHuang,LiLi,DongyangHuang,YunliYan,XiaonaDu,HailinZhang
Journal:Neuroscience(2016):356--367


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