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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/Cell Meter™ Fluorimetric Intracellular Nitric oxide (NO) Activity Assay Kit *Red Fluorescence Optimized f
产品编号:16356
市  场 价:¥100060.00
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:$5003.00
品      牌: AAT Bioquest
公      司:AAT Bioquest
公司分类:
AAT Bioquest/Cell Meter™ Fluorimetric Intracellular Nitric oxide (NO) Activity Assay Kit *Red Fluorescence Optimized f
商品介绍
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Ex/Em(nm)630/660
MWN/A
CAS#N/A
SolventN/A
StorageF/D/L
CategoryNeuroBIOLOGy
ReactiveOxygenSpecies
RelatedCellSignaling
BiochemicalAssays
Nitricoxide(NO)isanimportantbiologicalregulatorinvolvedinnumbersofphysiologicalandpathologicalprocesses.AlteredNOproductionisimplicatedinvariousimmunological,cardiovascular,neurodegenerativeandinflammatorydiseases.AsafreerADIcal,NOisrapidlyoxidizedandthereisrelativelylowconcentrationsofNOexistinginvivo.IthasbeenchallengingtodetectandunderstandtheroleofNOinbiologicalsystems.CellMeter™FluorimetricIntracellularNitricOxideAssayKitprovidesasensitivetooltomonitorintracellularNOlevelinlivecells.Nitrixyte™probesaredevelopedandusedinourkitasanexcellentreplacementforDAF-2forthedetectionandimagingoffreeNOincells.ComparedtothecommonlyusedDAF-2probe,Nitrixyte™probeshavebetterphotostABIlityandenhancedcellpermeability.ThisparticularkitusesNitrixyte™RedthatcanreactwithNOtogenerateabrightredfluorescentproductthathasspectralpropertiessimilartoTexasRed®.Nitrixyte™Redcanbereadilyloadedintolivecells,anditsfluorescencesignalcanbeconvenientlymonitoredusingthefiltersetofRed.Thiskitisoptimizedforflowcytometryapplications.
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Thisprotocolonlyprovidesaguideline,andshouldbemodifiedaccordingtoyourspecificneeds.

1.      Foreachsample,preparecellsin0.5mLwarmmediumorbufferofyourchoiceatadensityof5×105to1×106cells/mL.

Note:EachcelllineshouldbeevaluatedonanindividualbasistodeterminetheoptimalcelldensityforNOinduction.

 

2.       Add1μLof500XNitrixyte™Red(ComponentA)into0.5mLcellsUSPension. 

Note:Foradherentcells,gentlyliftthecellswith0.5mMEDTAtokeepthecellsintact,andwashthecellsoncewithserum-containingmediapriortoincubationwithNitrixyte™Red.

3.      IncubatecellswithtestcompoundsandNitrixyte™Red(fromStep2)at37ºCforadesiredperiodoftimetogenerateendogenousorexogenousNO.

Note1:Theappropriateincubationtimedependsontheindividualcelltypeandtestcompoundused.Optimizetheincubationtimeforeachexperiment.

Note2:WehaveusedRaw264.7cellsincubatedwithNitrixyte™Redworkingsolution,20µg/mLoflipopolysaccharide(LPS)and1mML-Arginine(L-Arg)incellculturemediumat37ºCfor16hours.

 

4.      ForaNONOatepositivecontroltreatment,add200uLofddH2OintothevialofComponentBtomake50mMstocksolution.Thestocksolutionisdilutedtomake1-2mMworkingsolutionwithAssayBuffer(ComponentC).Spindowncellsthathavepre-incubatedwithNitrixyte™Orangefor30minutes.Resuspendcellswith1mMDEANONOatepositivecontrolworkingsolution,andincubateat37ºCforanother30minutes.SeeFigure1fordetails.

 

5.      MonitorthefluorescenceintensityattheFL4channel(Ex/Em=630/660nm)usingaflowcytometer.Gateonthecellsofinterest,excludingdebris.

References&Citations
CitationExplorer

Fluorescentreal-timequantitativemeasurementsofintracellularperoxynitritegenerationandinhibition
Authors:ZhenLuo,QinZhao,JixiangLiu,JinfangLiao,RuoguPeng,YuntingXi,ZhenjunDiwu
Journal:AnalyticalBiochemistry(2017)

InducibleNitricOxideSynthase(iNOS)IsaNovelNegativeRegulatorofHematopoieticStem/ProgenitorCellTrafficking
Authors:MateuszAdamiak,AhmedABDelbaset-Ismail,JosephBMoore,JZhao,AhmedAbdel-Latif,MarcinWysoczynski,MariuszZRatajczak
Journal:StemCellReviewsandReports(2016):1--12


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