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1、1DevelopmentandCharacterizationofMouseHybridomasEugeneMechetnerSummaryCellfusionprotocolsthatweredevelopedbyKohlerandMilsteininthemid-1970sandaimedatproducingandcharacterizationofmousemonoclonalantibodies(MAbs)remainthegoldstandardofhybridomadevelopment.Despitetremendo
2、usprogressinusingMAbsinmultipleresearch,diagnostic,andtherapeuticareas,majorexperimentalflawsindesign-ingandcarryingouthybridomaexperimentationoftenresultintheproductionofhybrido-masexhibitingpoorgrowthparametersandsecretinglow-specificityandlow-affinityantibodies.This
3、methodologychapterisbuiltaroundtheconventionalhybridomaprotocol,withaspecialemphasisontissuecultureandbiochemicaltechniquesaimedatproducingtrulymonospecificandhighlyactivemouseMAbs.KeyWords:Hybridoma;monoclonalantibodies;MAbs;mouse;tissueculture.1.IntroductionTremendou
4、sprogressinthedevelopment,characterization,andmanufactur-ingofmonoclonalantibodies(MAbs)hasbeenmadesince1976,theyearwhenGeorgeJ.F.KohlerandCesarMilsteinpublishedtheirseminalpaper(1)ontheproductionofMAbsbyproducinghybridsbetweenmousesplenocyteswiththeirmyelomafusionpart
5、ner.Kohler’sandMilstein’soutstandingcontribution,forwhichtheywereawardedtogetherwithNielsK.Jernethe1984NobelPrizeinPhysiologyorMedicine,and—beyondall—theirdeliberate(and,alas,incom-prehensiblebytoday’sstandards)decisionnottopatentthehybridomatechnol-ogyresultedintherap
6、idandwidespreadadoptionofMAbsbybothacademiaandindustry.AsshowninTable1,overthelast30yrtwonewtypesofMAbs,recombinantandsynthetic,havebeendevelopedandvalidated.RecombinantMAbscanbeFrom:MethodsinMolecularBiology,vol.378:MonoclonalAntibodies:MethodsandProtocolsEditedby:M.A
7、lbitar©HumanaPressInc.,Totowa,NJ12MechetnerTable1ProgressinAntibodyDevelopmentSince197619762006AntibodytypesPolyclonalPolyclonalMonoclonalMonoclonal(animal)(animal)Monoclonal(recombinant)Monoclonal(synthetic)DiversityAminoacidsAminoacidsplatformNonaminoacid-based(e.g.,
8、aptamers)MonoclonalMouseMouseantibodies:RathostspeciesHamsterRabbitHumanChickenAlgaeExpressionMousecellsMouseandother
