AQC-solution-生命科学试剂-MCE

Hotline:400-820-3792Inhibitors•ScreeningLibraries•Proteinswww.MedChemEAQC(solution)Cat.No.:HY-DY1070CASNo.:148757-94-2分⼦式:C₁₄H₁₁N₃O₄分⼦量:285.26作⽤靶点:FluorescentDye作⽤通路:Others储存⽅式:PleasestoretheproductundertherecommendedconditionsintheCertificateofAnalysis.BIOLOGICALACTIVITY⽣物活性AQC(6-Aminoquinolyl-N-hydroxysccinimidylcarbamate)(solution)⼀种⾼效液相⾊谱荧光检测氨酸或蛋⽩质序列的试剂。AQC与⼀级和⼆级氨酸反应⽣成荧光衍⽣物，允许在⽪尔⽔平下检测氨酸。溶剂及浓度：DMSO：10mM体外研究Protocol1.ProteinPreparetion1)Inordertoobtainthebestlabelingeffect,pleasepreparetheprotein(antibody)concentrationas2mg/mL.2)ThepHvalueofproteinsolutionshallbe8.5±0.5.IfthepHislowerthan8.0,1Msodiumbicarbonateshallbeusedforadjustment.3)Iftheproteinconcentrationislowerthan2mg/mL,thelabelingefficiencywillbegreatlyreduced.Inordertoobtainthebestlabelingefficiency,itisrecommendedthatthefinalproteinconcentrationrangeis2-10mg/mL.4)Theproteinmustbeinthebufferwithoutprimaryamine(suchasTrisorglycine)andammoniumion,otherwisethelabelingefficiencywillbeaffected.2.CalculationofdyedosageTheamountofAQCrequiredforreactiondependsontheamountofproteintobelabeled,andtheoptimalmolarratioofAQCtoproteinisabout10.Example:assumingtherequiredmarkerproteinis500μL2mg/mLIgG(MW=150,000),use100μLDMSOdissolve1mgAQC,therequiredAQCvolumeis1.91μL,andthedetailedcalculationprocessisasfollows:1)mmol(IgG)=mg/mL(IgG)×mL(IgG)/MW(IgG)=2mg/mL×0.5mL/150,000mg/mmol=6.7×10-6mmol2)mmol(AQC)=mmol(IgG)×10=6.7×10-6mmol×10=6.7×10-5mmol3)μL(AQC)=mmol(AQC)×MW(AQC)/mg/μL(AQC)=6.7×10-5mmol×285.25mg/mmol/0.01mg/μL=1/2MasterofBioactiveMolecules—您⾝边的抑制剂⼤师www.MedChemE1.91μL(AQC)4.Runconjugationreaction1)Agoodvolumeoffreshlyprepared10mg/mLAQCisslowlyaddedto0.5mLproteinsampleInsolution,gentlyshaketomix,thencentrifugebrieflytocollectthesampleatthebottomofthereactiontube.Don'tmixwelltopreventproteinsamplesfromdenaturationandinactivation.2)Thereactiontubuleswereplacedinadarkplaceandincubatedgentlyatroomtemperaturefor60minutesatintervals.For10-15minutes,gentlyreversethereactiontubulesseveraltimestofullymixthetworeactantsandraisethebarefficiency.5.PurifytheconjugationThefollowingprotocolisanexampleofdye-proteinconjugatepurificationbyusingaSepHadexG-25column.1)PrepareSepHadexG-25columnaccordingtothemanufactureinstruction.2)Loadthereactionmixture(From"Runconjugationreaction")tothetopoftheSepHadexG-25column.3)AddPBS(pH7.2-7.4)assoonasthesamplerunsjustbelowthetopresinsurface.4)AddmorePBS(pH7.2-7.4)tothedesiredsampletocompletethecolumnpurification.Combinethefractionsthatcontainthedesireddye-proteinconjugate.REFERENCES[1].BoschL,etal.Applicationofthe6-aminoquinolyl-N-hydroxysccinimidylcarbamate(AQC)reagenttotheRP-HPLCdeterminationofaminoacidsininfantfoods.JChromatogrBAnalytTechnolBiomedLifeSci.2006;831(1-2):176-183.[2].CohenSA,etal.Applicationsofaminoacidderivatizationwith6-aminoquinolyl-N-hydroxysuccinimidylcarbamate.Analysisoffeedgrains,intravenoussolutionsandglycoproteins.JChromatogrA.1994;661(1-2):25-34.McePdfHeightCaution:Producthasnotbeen