德国莱卡刀片徕卡Leica818、819一次性刀片_
来自 : 蚂蚁淘
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| Author:NanciDonacki |
| Source:ContributedbyNanciDonacki |
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ReagentsandMaterials PBS PBS:0.05MEDTA PBS:0.01%thimerosal ConjugateStorageBuffer SATA DMF Hydroxylaminehydrochloride NaOHPellets PD10ColumnorSephadexG25 Sulfo-SMCC
PreparationofThiolatedAntibody Concentratetheantibodyto5-7mg/ml Dissolve10mgofSATAin1mlDMF(SATA:DMF) Add3.0mlSATA:DMFpermgofantibody Gentlystirsolutionfor2hoursatroomtemperature. Dissolve0.5ghydroxylaminehydrochloridein10mlPBS:EDTA Add0.25gNaOHpelletstothissolutiontoneutralizetoapproximatelypH7.0 Addtoantibodysolutionat6.49ml/mlSATA. Gentlystirfor30minutesatroomtemperature. EquilibratecolumnwithPBS:EDTA. Desaltthiolatedantibodyoncolumn.CollectinthesmallestvolumepossIBLewithoutcollectinganySATA.AllowfreeSATA(2ndpeak)tocomeoffthecolumn. Storethethiolatedantibodyat2-8oCfornomorethanonehour. Re-equilibratethecolumnwithPBS.
PreparationofMaleimide:HRP Weighout1.067mgHRPforeachmgofthiolatedantibody. Add1.1mgSulfo-SMCCpermgofHRP AddPBStoafinalconcentrationof8mg/mlHRP Mixfor2hoursonarotator. DesalttheHRPonPBSequilibratedcolumn.Collectthefirstpeak.
PreparationofAb:HRPConjugate ImmediatelymixtheHRPwiththethiolatedantibodyat1mgHRPpermgantibody. Gentlystirfor16-20hoursat2-8oC. Dialyzeagainst3changedofPBS,changingevery2hours,100Xtheconjugatevolume. Dilutetheconjugate2-foldinConjugateStorageBuffer.SterileFilter.
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