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1、抗11B轻类固醇脱氢酶1的单克隆抗体的制备与鉴定作者：王婉,杨金菊，刘莉，陈志成，高媛，高建恩，安立国，孙启鸿【摘要】冃的：制备抗人113拜类固醇脱氢酶1(homosapienshydroxysteroidllbetadehydrogenase1,HSD11B1)的单克隆抗休(mAb)并初步鉴定其特性。方法:将正常成人肝组织匀浆离心并分离线粒体，用线粒体总蛋白免疫BALB/c小鼠，采用杂交瘤技术制备mAb,并通过间接ELISA法、Westernblot、免疫组化的方法对mAb进行特性鉴定，通过免疫沉淀、UniZAPXR肝脏cDNA表达文库筛选鉴
2、定抗原。结果:通过间接ELISA筛选获得1株可稳定分泌抗人HSDllBlmAb的杂交瘤细胞系。其分泌的mAb的Tg亚类(型)为TgGl(k),Westernblot结果显示该mAb可以特异地识别相对分了质量(Mr)为35000的蛋白；应用mAbCBF245对人肝脏cDNA表达文库(UniZAPXR)进行筛选，阳性噬菌斑测序结果显示5个阳性克隆插入序列均为HSDllBlo结论:mAbBAD062特异地识别抗原HSD11B1,该mAb可用于ELTSA检测、Westernblot.免疫组化和免疫沉淀实验，为HSD11B1的研究提供了有力的工具。【关键
3、词】HSD11B1单克隆抗体线粒体[Abstract]ATM:Togenerateandidentifymonoclonalantibodies(mAb)againsthomosapienshydroxysteroidllbetadehydrogenase1(HSD11B1)・METHODS:NonnalhinnEinlivei'tissueswerehomogenized,andmitochondriawereisolatedbydifferentialcentrifugation.Thetotalmitochondrialproteinsw
4、ereusedtoimmunizeBALB/cmicetogeneratemAbbyhybridomatechnique・ThemAbwerecharacterizedbyELISA,Westernblotandimmunohistochemistry.Theantibodyspecificitywasidentifiedbyimmunoprecipitation(IP),andconfirmedbyUniZAPexpressionlibraryscreening・RESULTS:0nehybridomaCBF245secretingspeci
5、ficmAbagainstHSD1lBlwasestablished.TheTgsubclassofthismAbwasIgGl,anditcouldbeusedinELISA,Westernblot,immunohistochemistry.Oui'datashowedthattheantigenrecognizedbyCBF245mAbwaslocalizedinthehepatocytecytopiasm,withmolecularweightofMr35000inthemitochondriaofhumanlivertissue・The
6、CBF245mAbwasfurtherconfirmedbyimnmnoscreeningofUniZAPXRlivercDNAexpressionlibrary.CONCLUSION:Ahybridomace1IlinestablysecretingspecificniAbagainstHSDl1B1isestablishedandcharacterized・ThismAbreactedwithHSDl1B1inELISA,Westernblot,immunohistochemistryassay,IP,andwouldbeveryusefu
7、lfortheHSDllBlstudies.[Keywords]HSD11B1;mAb:mitoehondria11B径类固醇脱氢酶1(homosapienshydroxysteroidl1betadehydrogenasel,HSD11B1)属于短链脱氢/还原酶蛋白超家族(SDR),为相对分子质量(Mr)为34000的糖蛋白。此酶主要以糖基化的形式存在于微粒体，其作用依赖于辅酶NADP,在体内负责活性的糖皮质激素向其无活性前体的转换，调节局部糖皮质激素浓度，并决定糖皮质激素流向其受体［1］。代谢综合征糖皮质激素敏感性增加.HSD11B1调节异
8、常，可能是肥胖、糖耐量低减、高血压和心血管疾病等代谢综合征重要发病机制之一，选择性HSD11B1抑制剂有望成为其治疗靶点。HSD11B1在休内广泛表达
