请使用支持JavaScript的浏览器!
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/Amplite™ Fluorimetric Total NADP and NADPH Assay Kit *Red Fluorescence*/15259/400 Tests
产品编号:15259
市  场 价:¥85560.00
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:$4278.00
品      牌: AAT Bioquest
公      司:AAT Bioquest
公司分类:
AAT Bioquest/Amplite™ Fluorimetric Total NADP and NADPH Assay Kit *Red Fluorescence*/15259/400 Tests
商品介绍
Overview
PrinterFriendlyVersion

Ex/Em(nm)571/585
MWN/A
CAS#N/A
SolventN/A
StorageF/D/L
CategoryCellBIOLOGy
CellMetabolism
RelatedRedoxEnzymes
BiochemicalAssays
Nicotinamideadeninedinucleotide(NAD+)andnicotinamideadeninedinucleotidephosphate(NADP+)aretwoimportantcofactorsfoundincells.NADHisthereducedformofNAD+,andNAD+istheoxidizedformofNADH.ItformsNADPwiththeadditionofaphosphategrouptothe2"positionoftheadenylnucleotidethroughanesterlinkage.NADPisusedinanabolicbiologicalreactions,suchasfattyacidandnucleicacidsynthesis,whichrequireNADPHasareducingagent.Inchloroplasts,NADPisanoxidizingagentimportantinthepreliminaryreactionsofphotosynthesis.TheNADPHproducedbyphotosynthesisisthenusedasreducingpowerforthebiosyntheticreactionsintheCalvincycleofphotosynthesis.ThetrADItionalNAD/NADHandNADP/NADPHassaysaredonebymonitoringofNADHorNADPHabsorptionat340nm.ThismethodsufferslowsensitivityandhighinterferencesincetheassayisdoneintheUVrangethatrequiresexpensivequartzmicroplate.ThisAmplite™NADP/NADPHAssayKitprovidesaconvenientmethodforsensitivedetectionofNADPandNADPH.TheenzymesinthesystemspecificallyrecognizeNADP/NADPHinanenzymecyclingreaction.ThereisnoneedtopurifyNADP/NADPHfromsamplemix.Theenzymecyclingreactionsignificantlyincreasesdetectionsensitivity.Inaddition,thisassayhasverylowbackgroundsinceitisrunintheredvisIBLerangethatsignificantlyreducestheinterferencefrombiologicalsamples.Theassayhasdemonstratedhighsensitivityandlowinterferencewith570nmexcitation590nmemission.
SpectrumAdvancedSpectrumViewer



Thisprotocolonlyprovidesaguideline,andshouldbemodifiedaccordingtoyourspecificneeds.

1.PrepareNADPHstocksolution:

Add200µLofPBSbufferintothevialofNADPHStandard(ComponentC)tomake1mM(1nmol/µL)NADPHstocksolution.

Note:TheunusedNADPHstocksolutionshouldbedividedintosingleusealiquotsandstoredat-20oC.

 

2.PrepareNADP/NADPHreactionmixture:

Add10mLofNADP/NADPHSensorBuffer(ComponentB)intothebottleofNADP/NADPHRecyclingEnzymeMixture(ComponentA),andmixwell.

Note:ThisNADP/NADPHreactionmixtureisenoughfortwo96-wellplates.TheunusedNADP/NADPHreactionmixtureshouldbedividedintosingleusealiquotsandstoredat-20oC.

 

3.PrepareserialdilutionsofNADPHstandard(0to10μM):

3.1   Add10μLofNADPHstocksolution(fromStep1)to990µLPBSbuffertogenerate10µM(10 pmol/µL)NADPHstandardsolution.

Note:DilutedNADPHstandardsolutionisunstable,andshouldbeusedwithin4hours.

3.2   Take200μLof10μMNADPHstandardsolutiontoperform1:3serialdilutionstoget3,1,0.3,0.1,0.03,0.01,0.003and0μMserialdilutionsofNADPHstandard.

3.3   AddserialdilutionsofNADPHstandardandNADP/NADPHcontainingtestsamplesintoasolidblack96-wellmicroplateasdescribedinTables1and2.

Note:Preparecellsortissuesamplesasdesired.

Table1.LayoutofNADPHstandardsandtestsamplesinasolidblack96-wellmicroplate

BL

BL

TS

TS

….

….

 

 

 

 

 

 

NS1

NS1

….

….

….

….

 

 

 

 

 

 

NS2

NS2

 

 

 

 

 

 

 

 

 

 

NS3

NS3

 

 

 

 

 

 

 

 

 

 

NS4

NS4

 

 

 

 

 

 

 

 

 

 

NS5

NS5

 

 

 

 

 

 

 

 

 

 

NS6

NS6

 

 

 

 

 

 

 

 

 

 

NS7

NS7

 

 

 

 

 

 

 

 

 

 

Note:NS=NADPHStandards,BL=BlankControl,TS=TestSamples.

Table2.Reagentcompositionforeachwell

NADPHStandard

BlankControl

TestSample

SerialDilutions*:50μL

PBS:50μL

50μL

*Note:AddtheseriallydilutedNADPHstandardsfrom0.003μMto3μMintowellsfromNS1toNS7induplicate.HighconcentrationofNADPH(e.g.,>100μM,finalconcentration)maycausereducedfluorescencesignalduetotheoveroxidationofNADPHsensor(toanon-fluorescentproduct).

4.RunNADP/NADPHassayinsupernatants:

4.1   Add50μLofNADPHreactionmixture(fromStep2)intoeachwellofNADPHstandard,blankcontrol,andtestsamples(seeStep3.3)tomakethetotalNADPHassayvolumeof100µL/well

Note:Fora384-wellplate,add25μLofsampleand25μLofNADPHreactionmixtureintoeachwell.

4.2   Incubatethereactionatroomtemperaturefor15minutesto2hours,protectedfromlight.

4.3   MonitorthefluorescenceincreasewithafluorescenceplatereaderatEx/Em=530-570/590-600nm(optimalat540/590nm).

Note1:Thecontentsoftheplatecanalsobetransferredtoawhiteclearbottomplateandreadbyanabsorbancemicroplatereaderatthewavelengthof576±5nm.Theabsorptiondetectionhaslowersensitivitycomparedtofluorescencereading.

Note2:ForNADP/NADPHratiomeasurements,kit15264isrecommended.

Note3:ForcellbasedNADP/NADPHmeasurements,ReadiUse™mammaliancelllysisbuffer*5X*(cat#20012)isrecommendedtouseforlysingthecells.

References&Citations
CitationExplorer

CelastrolattenuatesangiotensinIImediatedhumanumbilicalveinendothelialcellsdamagethroughactivationofNrf2/ERK1/2/Nox2signalpathway
Authors:MiaoLi,XinLiu,YongpengHe,QingyinZheng,MinWang,YuWu,YuanpengZhang,ChaoyunWang
Journal:EuropeanJournalofPharmacology(2017):124--133

CytosolicRedoxStatusofWineYeast(SaccharomycesCerevisiae)underHyperosmoticStressduringIcewineFermentation
Authors:FeiYang,CaitlinHeit,DebraLInglis
Journal:Fermentation(2017):61

EpigeneticregulationofRunx2transcriptionandosteoblastdifferentiationbynicotinamidephosphoribosyltransferase
Authors:MinLing,PeixinHuang,ShamimaIslam,DanielPHeruth,XuananLi,LiQinZhang,Ding-YouLi,ZhaohuiHu,ShuiQingYe
Journal:Cell&Bioscience(2017):27

MCU-dependentmitochondrialCa2+inhibitsNAD+/SIRT3/SOD2pathwaytopromoteROSproductionandmetastasisofHCCcells
Authors:TRen,HZhang,JWang,JZhu,MJin,YWu,XGuo,LJi,QHuang,HYang
Journal:Oncogene(2017)

Metabolicandmolecularinsightsintoanessentialroleofnicotinamidephosphoribosyltransferase
Authors:LiQZhang,LeonVanHaandel,MinXiong,PeixinHuang,DanielPHeruth,CharlieBi,RogerGaedigk,XunJiang,Ding-YouLi,GeraldWyckoff
Journal:CellDeath&Disease(2017):e2705

PyrroloquinolineQuinone,aRedox-activeo-Quinone,StimulatesMitochondrialBiogenesisbyActivatingSIRT1/PGC-1αSignalingPathway
Authors:KazuhiroSaihara,RyosukeKamikubo,KazutoIkemoto,KojiUchida,MitsuguAkagawa
Journal:Biochemistry(2017)

ResveratrolattenuatesexcessiveethanolexposureinducedinsulinresistanceinratsviaimprovingNAD+/NADHratio
Authors:GangLuo,BingqingHuang,XiangQiu,LinXiao,NingWang,QinGao,WeiYang,LipingHao
Journal:MolecularNutrition&FoodResearch(2017)

ASnapshotofthePlantGlycatedProteomeSTRUCTURAL,FUNCTIONAL,ANDMECHANISTICASPECTS
Authors:TatianaBilova,ElenaLukasheva,DominicBrauch,UtaGreifenhagen,GaganPaudel,ElenaTarakhovskaya,NadezhdaFrolova,JulianeMittasch,GerdUlrichBalcke,AlainTissier
Journal:JournalofBiologicalChemistry(2016):7621--7636

AMPKactivationprotectscellsfromoxidativestress-inducedsenescenceviaautophagicfluxrestorationandintracellularNAD+elevation
Authors:XiaojuanHan,HaoranTai,XiaoboWang,ZheWang,JiaoZhou,XiaweiWei,YiDing,HuiGong,ChunfenMo,JieZhang
Journal:Agingcell(2016):416--427

Cell-LineSelectivityImprovesthePredictivePowerofPharmacogenomicAnalysesandHelpsIdentifyNADPHasBioMarkerforFerroptosisSensitivity
Authors:KenichiShimada,MikiHayano,NenCPagano,BrentRStockwell
Journal:Cellchemicalbiology(2016):225--235


ViewMoreCitations
品牌介绍

AAT Bioquest常用产品报价单

产品名货号公司分类商城分类美金价
AAT Bioquest/Fluorescein, disodium salt *CAS 518-47-8*/2/100 mg2iFluor Rapid Tests酸碱缓冲液1088.00
AAT Bioquest/14-3-3 ζ (Ab-58) Antibody/8B0001/50 ug8B0001Signaling Intermediate Ab重组抗体2828.00
AAT Bioquest/Opioid Receptor (Phospho-Ser375) Antibody/8A0022/50 ug8A0022Phospho-Specific Ab受体2828.00
AAT Bioquest/iFluor™ 350 goat anti-mouse IgG (H+L)/16440/200 ug16440Fluorescent Anti-IgGs荧光染料1088.00
AAT Bioquest/trFluor™ Eu goat anti-mouse IgG (H+L)/16518/100 ug16518Fluorescent Anti-IgGs抗小鼠4278.00
AAT Bioquest/RPE-streptavidin conjugate/16900/100 ug16900Streptavidin Conjugates其他生物染料1378.00
AAT Bioquest/GCNT3 Antibody/8C14708/50 ug8C14708Signaling Intermediate Ab功能性抗体2828.00
AAT Bioquest/Amplite™ Luciferase Reporter Gene Assay Kit *Bright Glow*/12518/1 plate12518Reporter Gene Enzymes其它检测试剂盒2103.00
AAT Bioquest/ReadiLink™ Rapid mFluor™ Violet 450 Antibody Labeling Kit *Microscale Optimized for Labeling 50 &mi1100mFluor™ Dyes and Kits标记试剂盒2103.00
AAT Bioquest/Amplite™ Colorimetric Urea Quantitation Kit *Blue Color*/10058/200 Tests10058Diagnostic Molecules定量试剂盒2828.00
AAT Bioquest/Screen Quest™ CHO-Gqi Chimera Cell line/38101/Each38101cAMP GPCR Assays常规细胞株0.00
自营商城图标
厂家直采
全球直采 正品优价
正品保障图标
正品保障
厂家直发 有线跟踪
解放采购图标
正规清关
CIF100%正规报关,提供发票
及时交付图标
及时交付
限时必达 不达必赔