ProductName | 20SProteasomeActivityKit |
Description | Fluorometricdetectionofpurified20Sproteasomeactivity |
SpeciesReactivity | SpeciesIndependent |
Platform | Microplate |
SampleTypes | PurifiedProteins |
DetectionMethod | FluorometricAssay |
AssayType | ContinuousKineticAssay |
Utility | Activitykitusedtoscreenpotentialproteasomeinhibitorsandactivators,andtomeasureandquantify20Sproteasomeactivityincontinuouskineticandend-pointassays. |
IncubationTime | 15-30minutes |
NumberofSamples | 40samplesinduplicateusingtheplateformat,or100x50µLmicro-cuvettebasedassays |
OtherResources | KitBooklet,MSDS |
StorageTemperature | 4ºC | |||||||||||||||||||||
ShippingTemperature | BlueIce | |||||||||||||||||||||
ProductType | ActivityKits | |||||||||||||||||||||
AssayOverview | The20SProteasomeActivitykitfacilitatestherapid,robustmeasurementofproteasomeactivityinvitro.Thekitutiliseshighpurity,fluorogenicsubstrateSuc-LLVY-AMCtogetherwithsuitablecalibrationstandardsandcontrolsfortheaccurateandsensitiveassessmentofproteasomechymotrypsin-like(β5)activity.Continuouskineticorend-pointassayscanbeperformedinmicro-cuvettesorin96-wellplateformatformulti-sampleanalysis.Containssufficientmaterialsforonefull96-wellplateassayorupto100x50µLmicro-cuvettebasedassaystoberun. | |||||||||||||||||||||
KitOverview |
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CiteThisProduct | 20SProteasomeActivityKit(StressMarqBiosciencesInc.,VictoriaBCCANADA,Catalog#SKT-133) |
AlternativeNames | Chymotrypsin-like(β5)ActivityKit |
ResearchAreas | Cancer,Apoptosis,CardiovascularSystem,CellSignaling,Neurodegeneration,Neuroscience,Post-translationalModifications |
ScientificBackground | Proteasomesarenon-lysosomalproteolyticcomplexeslocalisedprimarilyinthecytoplasmandinthenucleusofeukaryoticcells.TheyareresponsIBLefortheubiquitin-mediateddegradationofshorthalf-lifeproteinsandpeptidesthatareinvolvedinessentialcellularprocessesincludingcell-cycleregulation,apoptosisandtranscriptionalregulation,innateimmunityandantigenprocessing,andintheremovalofredundantordamagedproteins.Assuchproteindegradationbytheubiquitin-proteasomepathwayhasamajorregulatoryfunctionforproliferationactivityandsurvivalofbothnormalandmalignantcells,anditsdysfunctionhasbeenimplicatedinawiderangeofotherdiseaseprocessesincludingneurodegenerative,cardiovascularandmetabolicdisorders.The26Sproteasomestructureiscomposedofa20Sproteasomecatalyticcorecomplexandoneortwo19S(PA700)regulatorysubcomplexes.The20Scorecomprisestwocopiesof14subunits(7alphasubunitsand7betasubunits)arrangedinaα7β7β7α7cylindricalarray.Proteolyticactivitiesaredeterminedbytheβ1(caspase-like),β2(trypsin-like)andβ5(chymotrypsin-like)subunits,accesstowhichisguardedbytheα-subunits.The19SregulatoryunitconsistsofsixATPaseandatleasttennon-ATPasesubunitsthatarerequiredforubiquitinatedproteinbinding,deubiquitination,substrateunfoldingandtranslocationtothe20Scatalyticcore.Varyingcatalyticsubunitcomposition(β1,β1i;β2,β2i;β5,β5i)resultsinavarietyofpossiblesubtypesfromfullconstitutiveproteasome(β1,β2,β5)throughmixedpopulationstofullinducible/immunoproteasome(β1i,β2i,β5i).AlternativeregulatorycomplexessuchasthePA200and11Sproteasomeactivatorsconferdifferentsubstratespecificitiesandactivitycomparedtothe19Sregulator. |
References | 1.Saeki,Y.&Tanaka,K.Methodsinmolecularbiology(Clifton,N.J.)832,315–37(2012). 2.Frankland-Searby,S.&Bhaumik,S.R.Biochimicaetbiophysicaacta1825,64–76(2012). 3.Basler,M.,Kirk,C.J.&Groettrup,M.Currentopinioninimmunology1–7(2012). 4.Löw,P.Generalandcomparativeendocrinology172,39–43(2011). 5.Dennissen,F.J.a,Kholod,N.&VanLeeuwen,F.W.Progressinneurobiology96,190–207(2012). 6.Geng,F.,Wenzel,S.&Tansey,W.P.Annualreviewofbiochemistry81,177–201(2012). |
AMCStandardCurvefor20SProteasomeActivityKit.Representativeplotoffluorescencemeasurements(RFU)ofserialdilutionsofAMCStandardatrecommendedconcentrations.
Continuouskineticenzymeactivityassayshowingthetimecourseofcontrol20SproteasomeactivitywithSuc-LLVY-AMCsubstrateusingtheStressXpress®20SProteasomeActivityKit.20Sproteasome(~2.5nM)wasincubatedwith100µMSuc-LLVY-AMCinproteasomeassaybufferfor30minutesatroomtemperaturealongsideasubstrateonly(blank)controlreaction.Fluorescencemeasurements(RFU)weretakenat30secondintervalsandplottedvs.time.
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StressMarq Biosciences Inc.位于加拿大维多利亚,是一家生物科技公司,专门从事试剂与试剂盒研究。我们有强大的国际经销商网络,主要为私人和公众客户提供经我们多次试验成功的试剂,服务范围遍及全球40多个国家。 我们公司的核心技术领域为细胞应激与离子通道以及载体研究,同时在其他领域也取得了一定成就,包括翻译后修饰,提供甲基化与乙酰基化抗体。其中,细胞应激领域主要包括热休克蛋白(HSP)领域。我们公司不仅在热休克蛋白领域领先全球,而且在氧化应激领域也卓有成就。StressMarq的优势在于提供四种独立的产品系列,分别涉及抗体、蛋白、酶联免疫吸附试验(ELISA)试剂盒及小分子领域。 通过定位好以上四个领域,研究人员能够运用研究相关的整套试剂工具,将研究产品重点放在热休克蛋白70和90(HSP70和HSP90)两个核心领域。产品主要包括ELISA试剂盒、抑制剂、蛋白质(包括无内毒素制剂,而非低内毒素制剂)以及经过高级验证的抗体,产品在全球范围内均有销售。 StressMarq因其试剂验证质量过硬,跻身为高端公司。研究人员能放心使用抗体产品,因为本产品将具有免疫印迹(WB)、免疫沉淀、组织染色技术的功能,如免疫组织化学(IHC)和免疫细胞化学(ICC)。 本公司提供氧化应激产品,如8OHdG(DNA受损)ELISA试剂盒,同时还提供专门的抗体,适用于非定量技术,如免疫组化。在免疫组化中,组织的目标可视化更为重要。除了提供用于进一步研究的相关工具外,StressMarq还提供新的尖端研究工具,用于开发新的生物项目,包括全球独一无二的α B晶体蛋白ELISA试剂盒(现在是公认的乳腺癌标志物)以及热休克同源蛋白70(Hsc70)ELISA试剂盒。本公司已将专业技术扩展到抗体、小分子离子通道与载体等更新领域,前者包括当前热门的核心领域:Nav、Cav、TRP、Kv、KCNQ、HCN,后者包括当前热门的核心领域:ENaC、NCC和NKCC2以及主要水通道蛋白载体。