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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/Amplite™ Colorimetric Urea Quantitation Kit *Blue Color*/10058/200 Tests
产品编号:10058
市  场 价:¥56560.00
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:$2828.00
品      牌: AAT Bioquest
公      司:AAT Bioquest
公司分类:
AAT Bioquest/Amplite™ Colorimetric Urea Quantitation Kit *Blue Color*/10058/200 Tests
商品介绍
Overview
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Ex/Em(nm)650/None
MWN/A
CAS#N/A
SolventN/A
StorageF/D/L
CategorySmallMoleculeDetection
DiagnosticMolecules
RelatedCellMetabolism
BiochemicalAssays
Ureaisthefinaldegradationproductofproteinandaminoacidmetabolisminanimals.Itisproducedinliver,secretedbykidneyandexcretedthroughurine.Thedeterminationofureaisveryusefultestinclinicallaboratorytomonitorhealthstatus.TheBloodUreaNitrogen(BUN)testisameasureoftheamountofnitrogeninthebloodintheformofureaandisprimarilyused,alongwiththecreatininetesttoevaluatekidneyfunction,helpingdiagnosekidneydiseases.OurAmplite™ColorimetricUreaAssayKitprovidesasimpleandsensitivecolorimetricmethodforthequantitationofureaconcentrationinBIOLOGicalsamplessuchasserum,plasmaandurine,etc.Theassayisbasedonanenzyme-coupledreactionofureaintheassaybuffer,andfinallyproducesabluecoloredproduct.Theintensityofcolorproducedisproportionaltotheconcentrationofureainthesample,whichcanbemeasuredcolorimetricallyat660-670nm.ThisAmplite™ColorimetricUreaAssayKitprovidesasimpleassaytodetectaslittleas10µMureaina150µLassayvolume.Theassaycanbeperformedinaconvenient96-wellor384-wellmicrotiter-plateformatandeasilyadaptedtoautomationwithoutaseparationstep.

Thisprotocolonlyprovidesaguideline,andshouldbemodifiedaccordingtoyourspecificneeds.

1.      Prepareserialdilutionsofurea(0to1mM)solutions:

 

1.1   Add1μLof1.0MUreaStandard(ComponentD)to999µLDPBStogenerate1.0mMstandardureasolution.

 

1.2   Take300μLof1.0mMstandardtoperform1:3serialdilutionstoget300,100,30,10,3,1,and0µMstandardureasolutions.

 

1.3   Addureastandardsandureacontainingtestsamplesintoa96-wellclearbottommicroplateasdescribedinTables1and2.

 

Table1.Layoutofureastandardsandtestsamplesinaclearbottom96-wellmicroplate:

BL

BL

TS

TS

….

….

 

 

 

 

 

 

US1

US1

….

….

….

….

 

 

 

 

 

 

US2

US2

 

 

 

 

 

 

 

 

 

 

US3

US3

 

 

 

 

 

 

 

 

 

 

US4

US4

 

 

 

 

 

 

 

 

 

 

US5

US5

 

 

 

 

 

 

 

 

 

 

US6

US6

 

 

 

 

 

 

 

 

 

 

US7

US7

 

 

 

 

 

 

 

 

 

 

       Note:US=UreaStandards,BL=BlankControl,TS=TestSamples.

 

Table2.Reagentcompositionforeachwell:

UreaStandard

BlankControl

TestSample

Serialdilutions*:50μL

DPBS:50μL

50μL

*Note:Addtheseriallydilutedureastandardsfrom1to1000µMintowellsfromUS1toUS7induplicate.

 

2.      PrepareAssayReactionMixtureI:

2.1   Add100µLofddH2OintothevialofAssayEnzymeMix(ComponentA)tomake100XAssayEnzymeMixsolution.

 

2.2   Add50μLreconstitutedAssayEnzymeMixsolutioninto5mLAssayBufferItomakeAssayReactionMixtureI.

Note1:TheAssayreactionmixtureIshouldbeusedpromptlyandkeptfromlight.Theassaysensitivitywillbedecreasedwithlongerstoragetime.ThefreshAssayEnzymeMixtureIisrecommendedforthebestresult.

Note2:Anyremaining100Xenzymemixsolution(fromStep2.1)needstobealiquotandfrozenat-20°C,avoidrepeatedfreeze-thawcycles.

 

3.      Runureaassay:

3.1   Add50μLofAssayReactionMixtureI(fromStep2.2)toeachwelloftheureastandard,blankcontrol,andtestsamples(seeStep1.3)sothatthetotalassayvolumeis100µL/well.

Note:Fora384-wellplate,add25μLsample,25μLofassayreactionmixtureIperwell.

3.2   Incubatethereactionfor30-60minutesatroomtemperatureor37°C,protectedfromlight.

3.3   Add50μLofAssayBufferII(ComponentC)toeachwellsothatthetotalassayvolumeis150µL/well.

Note:Fora384-wellplate,add25μLAssayBufferII(ComponentC)toeachwell.

3.4   Incubateatroomtemperaturefor10-15minutes,andmonitortheabsorbanceincreaseat660-670nmusinganabsorbancemicroplatereader.

Note1:ThecolorturnstoyellowafterAssayBufferII(ComponentC)isadded,andthewellswithureastandardorsampleswillshowblue-greencolorafterincubation.Theintensityofthecolorwillreachthemaximumin15-30minutes,andisproportionaltotheconcentrationofurea.

Note2:Thefinalcolorisstablefor~1hourinroomtemperatureandthecolorintensitywilldecreasewithtime.

References&Citations
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1.   LawsonTS,ConnallyRE,VemulpadS,PiperJA.(2011)Dimethylformamide-free,urea-NaClfluorescenceinsituhybridizationassayforStaphylococcusaureus.LettApplMicrobiol.

2.   LiangA,QinH,ZhouL,ZhangY,OuyangH,WangP,JiangZ.(2011)AnovelandsensitiveresonancescatteringassayfordetectionofureainserumcoupledureasecatalyticreactionandNH4+associatedparticlereaction.BioprocessBiosystEng,34,639.

3.   FerrucioB,FranchiCA,BoldrinNF,deOliveiraML,deCamargoJL.(2010)Evaluationofdiuron(3-[3,4-dichlorophenyl]-1,1-dimethylurea)inatwo-stagemouseskincarcinogenesisassay.ToxicolPathol,38,756.

4.   NgokaLC.(2008)Sampleprepforproteomicsofbreastcancer:proteomicsandgeneontologyrevealdramaticdifferencesinproteinsolubilizationpreferencesofrADIoimmunoprecipitationassayandurealysisbuffers.ProteomeSci,6,30.

5.   MulagalapalliS,KumarS,KalathurRC,KayasthaAM.(2007)Immobilizationofureasefrompigeonpea(Cajanuscajan)onagartabletsanditsapplicationinureaassay.ApplBiochemBiotechnol,142,291.

6.   RozetE,WascotteV,LecouturierN,PreatV,DeweW,BoulangerB,HubertP.(2007)ImprovementofthedecisionefficiencyoftheaccuracyprofilebymeansofadesirABIlityfunctionforanalyticalmethodsvalidation.Applicationtoadiacetyl-monoximecolorimetricassayusedforthedeterminationofureaintransdermaliontophoreticextracts.AnalChimActa,591,239.

7.   KimuraS,IyamaS,YamaguchiY,KanakuraY.(2003)Newenzymaticassayforserumureanitrogenusingureaamidolyase.JClinLabAnal,17,52.

8.   FarkasA,VamosR,BajorT,MullnerN,LazarA,HrabaA.(2003)Utilizationoflacrimalureaassayinthemonitoringofhemodialysis:conditions,limitationsandlacrimalarginasecharacterization.ExpEyeRes,76,183.

9.   KimuraS,YamanishiH,IyamaS,YamaguchiY,KanakuraY.(2003)Enzymaticassayfordeterminationofbicarbonateioninplasmausingureaamidolyase.ClinChimActa,328,179.

10.   BilardiC,BiaginiR,DulbeccoP,IiritanoE,GambaroC,MeleMR,BorroP,TessieriL,ZentilinP,MansiC,VigneriS,SavarinoV.(2002)Stoolantigenassay(HpSA)islessreliablethanureabreathtestforpost-treatmentdiagnosisofHelicobacterpyloriinfection.AlimentPharmacolTher,16,1733.


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