DEAE-Sepharose-Fast-Flow-说明书

Instructions71-5009-64AFIonexchangemediaCMSepharose™FastFlowDEAESepharoseFastFlowQSepharoseFastFlowSPSepharoseFastFlowCM,DEAE,QandSPSepharoseFastFlowionexchangersarepartoftheBioProcess™mediaproductportfolio.Thesechromatographymedia(resins)reflectstheimportantroletheyplayinproteinpurificationtoday.Thereliabilityandwelldocumentedperformanceofthesemedia,havemadethemacommonchoiceforcaptureandintermediatepurificationofproteinsinbothresearchandindustry.SepharoseFastFlowionexchangersoffermanypracticaladvantages:•Highbindingcapacityandgoodflowproperties•Highchemicalandphysicalstabilitiesincombinationwithpredictablescale-up•Reliableandreproducibleperformance•Easyandeffectivecleaning-in-place(CIP)/sanitization•Various,convenientprepackedcolumnformats•SecurityofsupplyandcomprehensiveregulatorysupportTableofContents31BioProcesschromatographymedia................................................32CharacteristicsofSepharoseFastFlowionexchangers........123Methodoptimization...............................................................................144Scaleup.........................................................................................................165Packingcolumns.......................................................................................206Evaluationofcolumnpacking............................................................227Maintenance...............................................................................................258Orderinginformation..............................................................................Pleasereadtheseinstructionscarefullybeforeusingtheproducts.SafetyForuseandhandlingoftheproductsinasafeway,pleaserefertotheSafetyDataSheets.271-5009-64AF1BioProcesschromatographymediaBioProcesschromatographymediaaredevelopedandsupportedforproductionscalechromatography.AllBioProcesschromatographymediaareproducedwithvalidatedmethodsandaretestedtomeetmanufacturingrequirements.Secureorderinganddeliveryroutinesgiveareliablesupplyofchromatographymediaforproductionscale.RegulatorySupportFiles(RSF)areavailabletoassistprocessvalidationandsubmissionstoregulatoryauthorities.BioProcessmediacoverallpurificationstepsfromcapturetopolishing.2CharacteristicsofSepharoseFastFlowionexchangersIntroductionThebasematrixofSepharoseFastFlowionexchangersishighlycross-linkedagarosewhichgivestheionexchangershighchemicalandphysicalstability.Thismeansthatcharacteristicssuchascapacity,elutionbehaviorandpressure/flowrateareunaffectedbythesolutionscommonlyusedinprocesschromatographyandcleaningprocedures,fordetailsseetableundereachrespectiveionexchanger.Highphysicalstabilitygivesgoodflowcharacteristicsandlowbackpressures.Flowvelocitiesrangingbetween300and700cm/hthroughabedheightof15cmatapressureof1bararetypicalforthesemedia,seeFigure1.Furthermore,thehighrigidityofthematrixminimizesvolumevariationsduringchangeofpHorionicstrength.71-5009-64AF3Linearflowrate(cm/h)700600500400300200100123P(bar)Fig1.Atypicalpressure/flowratecurveforSepharoseFastFlowionexchangers.471-5009-64AFCharacteristicsofCMSepharoseFastFlowCMSepharoseFastFlowisaweakcationexchanger.Theionexchangegroupisacarboxymethylgroup,seebelow.–O–CH2COO-Table1.CharacteristicsofCMSepharoseFastFlow.DescriptionPropertyWeakcationIonexchangetype0.09to0.13mmol/mlmediumTotalioniccapacityCross-linkedagarose,6%Matrix90μmAverageparticlesize(d50v)1300to600cm/h(15cmbedheight,1bar,25°C,XK50/30column)Flowvelocity4°Cto40°CWorkingtemperature2to14(Cleaning-in-place)4to13(Workingrange)pHstability2Allcommonlyusedaqueousbuffers1MNaOH,8Murea,6Mguanidinehydrochloride,70%ethanolChemicalstabilityOxidizingagents,cationicdetergentsLongexposures(1week,20°C)topH4Avoid4°Cto30°CStoragetemperature20%ethanolStoragebuffer31d50visthemedianparticlesizeofthecumulativevolumedistribution.2Workingrange:pHintervalwherethemediumcanbeoperatedwithoutsignificantchangeinfunction.Cleaning-in-place:pHstabilitywherethemediumcanbesubjectedtocleaning-in-placewithoutsignificantchangeinfunction.3LargepacksizesofCMSepharoseFastFlowin2%benzylalcoholareavailableonrequest.ContactyourlocalGErepresentativeforfurtherinformation.ThetitrationcurveinFigure2showsthepHworkingrangeofCMSepharoseFastFlow,i.e.,thepHrangeinwhichtheCMgroupischarged.71-5009-64AF5CMSepharoseFastFlowpH12108642100200300400500600700800ml0.1MNaOHFig2.TitrationcurveofCMSepharoseFastFlow.671-5009-64AFCharacteristicsofDEAESepharoseFastFlowDEAESepharoseFastFlowisaweakanionexchanger.Theionexchangegroupisadiethylaminoethylgroup,seebelow.–O–CH2CH2-N+(C2H5)2HTable2.CharacteristicsofDEAESepharoseFastFlow.DescriptionPropertyWeakanionIonexchangetype0.11to0.16mmol/mlmediumTotalioniccapacityCross-linkedagarose,6%Matrix90μmAverageparticlesize(d50v)1300to600cm/h(15cmbedheight,1bar,25°C,XK50/30column)Flowvelocity4°Cto40°CWorkingtemperature2to14(Cleaning-in-place)2to12(Workingrange)pHstability2Allcommonlyusedaqueousbuffers1MNaOH,8Murea,6Mguanidinehydrochloride,70%ethanolChemicalstabilityOxidizingagents,anionicdetergentsLongexposures(1week,20°C)topH4Avoid4°Cto