CypK-N-Cyclopropene-L-Lysine-DataSheet-生命科学试剂-MedChemExpress

Hotline:400-820-3792Inhibitors•ScreeningLibraries•Proteinswww.MedChemECypKCat.No.:HY-134669CASNo.:1610703-09-7Synonyms:N-Cyclopropene-L-Lysine分⼦式:C₁₂H₂₀N₂O₄分⼦量:256.3作⽤靶点:Others作⽤通路:Others储存⽅式:PleasestoretheproductundertherecommendedconditionsintheCertificateofAnalysis.BIOLOGICALACTIVITY⽣物活性CypK(N-Cyclopropene-L-Lysine)赖氨酸的环丙烯衍⽣物，通过因编码扩增有效地整合到抗体中。CypK⼀个最⼩的⽣物正交柄，⽤于创建稳定的治疗蛋⽩偶联物。体外研究Guidelines(Followingisourrecommendedprotocol.Thisprotocolonlyprovidesaguideline,andshouldbemodifiedaccordingtoyourspecificneeds).CypKassay[1](expresstheantibody):1.ThawavialofHEKsuspensioncellsina250mLflaskcontaining50mLofexpressionmediumsupplementedwith100units/mLpenicillin,100µg/mLstreptomycin,and250ng/mLamphotericinB.Keepthecellsat37oCwith8%CO2inhumidifiedincubatorsequippedwithashakerat125rpm.Splitcellsto0.3-0.5x10[6]cells/mL(every2-3days)atleast2timesbeforetransfecting.2.Whenadensityof2.5x10[6]cells/mLisreached(2-3daysaftersplitting),prepareafreshsolutionof100mMCypK.Forthispurpose,weigh64mgofCypK,add2.5mLof0.1sodiumhydroxide,vortex,spindowntorecoverallundissolvedparticlesandsonicate.3.Add2.5mLofCypK(100mMin0.1MNaOH)to42.5mLofexpressionmediumsupplementedwithantibiotics.Mixwell,add250µLof0.1MHCl,andsterilizeusinga0.22µmfilter.4.Dilute50µgofHCandLCpKym1plasmidsto2.5mLwithreducedserummedium.Inaseparatetube,dilute135µLoftransfectionreagentto2.5mLwithreducedserummedium.5.Fiveminutesafterpreparingthesolutions,mixtheplasmidsandthetransfectionreagentsolutionandincubatefor20mintoallowtheformationofcomplexesbetweentheDNAandthetransfectionreagent.6.Inthemeantime,centrifuge125millioncellsatthetargetdensityfor5minat500xg,resuspendwiththeexpressionmediumcontainingCypKandaddtheDNA–transfectionreagentmixture.1/2MasterofBioactiveMolecules—您⾝边的抑制剂⼤师www.MedChemE7.Afterincubatingcellsfor20h,add250µLoftransfectionreagentenhancersincludedinthekit.8.Harvestantibodiesfromthesupernatant6-7daysafteradditionofCypK(nochangeofmediumisrequiredduringexpression).REFERENCES[1].Oller-SalviaB.GeneticEncodingofaNon-CanonicalAminoAcidfortheGenerationofAntibody-DrugConjugatesThroughaFastBioorthogonalReaction.JVisExp.2018Sep14;(139):58066.[2].Oller-SalviaB,et,al.RapidandEfficientGenerationofStableAntibody-DrugConjugatesviaanEncodedCyclopropeneandanInverse-Electron-DemandDiels-AlderReaction.AngewChemIntEdEngl.2018Mar5;57(11):2831-2834.McePdfHeightCaution:Producthasnotbeenfullyvalidatedformedicalappli