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1、中华神经外科疾病研究杂志(ChinJNeurosurgDisRes)2005;4(3)#245#文章编号:1671-2897(2005)04-245-03#论著#人脑胶质瘤细胞的原代培养及生长活性观察易伟牛洪泉陶胜忠叶飞雷霆薛德麟(华中科技大学同济医学院附属同济医院神经外科,湖北武汉430030)摘要目的研究人脑胶质瘤细胞原代培养的方法,观察培养细胞的生长活性与肿瘤级别之间的关系。方法用消化培养法原代培养24例人脑胶质瘤细胞,酶消化法+振荡贴壁法+机械划除法纯化,增殖细胞核抗原(PCNA)免疫组化染色及甲基噻唑基四唑(MTT)法观察培养细胞的生长活性。结果
2、20例原代培养成功,第4代以后肿瘤细胞纯度达98%以上,PCNA、MTT法显示肿瘤细胞的生长活性与肿瘤级别成正比。结论采用消化培养法可成功培养原代胶质瘤细胞,酶消化法+振荡贴壁法+机械划除法的纯化方法可获得较高的肿瘤细胞纯度;肿瘤细胞的生长活性与肿瘤的恶性程度有关,离体细胞的生长活性可作为判断肿瘤恶性程度的一个指标;检测细胞生长活性MTT法比PCNA染色更具敏感性。关键词胶质瘤;原代培养;增殖细胞核抗原(PCNA);MTT试验中国图书资料分类号R329.2文献标识码ACharacterizationandgrowthactivityobservationo
3、fprimaryhumangliomascellcultureYIWei,NIUHongquan,TAOShengzhong,YEFei,LEITing,XUEDelinDepartmentofNeurosurgery,TongjiHospital,TongjiMedicalCollege,HuazhongUniversityofScienceandTechnology,Wuhan430030,ChinaAbstractObjectiveToinvestigatethemethodsofprmiaryhumangliomascellcultureandth
4、erelationshipbetweentheculturedcellsandtheirhistologicalgrades.MethodsTwenty-fourprmiarygliomaswereculturedwithenzymaticdigestionandthetumorcellswerepurifiedwithanewprotoco.lTheprocesswasdigestingthecellswithtrypsinatfirs,tthenoscillatingthecellswhiletheyadhered,andmechanicallyscr
5、apingthemipur-itycellsoffifnecessaryatfina.lInordertoobservethecells'proliferation,methylthiagolyltetragolium(MTT)assayandproliferatingcellnuclearantigen(PCNA)mimunohistochemicalstainingwereapplied.ResultsTwentyprmiarygliomasweresuccessfullycultured.Thenewpurificationprotocolcould
6、purifythetumourcellsbeyond98%.PCNAandMTTassaydemonstratedtheproliferationoftheculturedcellscorrelatedwiththeirhistologicalgrades.ConclusionThemethodofdigestioncultivationcansuccessfullycultureprmiarygliomas.Usingthepur-ificationmethodofdigestion-oscillation-scrapecanobtainahighpur
7、ityoftumorcells.Proliferationofculturedpr-imarytumorcellscanbeanindicatorofthetumors'biologicalmalignancy.MTTassayismoresensitivethanPCNAmimunohistochemicalstainingforobservingcells'proliferation.KeywordsGlioma;Prmiarycellculture;ProliferatingCellNuclearAntigen(PCNA);(MTT)assay细胞培
8、养已成为近年来最为重要的研究方法之(proliferatingce
