cd59特异位点的短肽封条对hela细胞凋亡作用的研究

《cd59特异位点的短肽封条对hela细胞凋亡作用的研究》由会员上传分享，免费在线阅读，更多相关内容在学术论文-天天文库。

1、CD59特异位点的短肽封条对HeLa细胞凋亡作用的研究作者：李丽,池沛冬,孟锐,杨滨燕,吴长有【关键词】BrdU增殖流式细胞仪细胞活化　　[Abstract]AIM:ToinvestigatethecorrelationofBrdUincorporationwiththeactivationandcytokineexpressionofTcells.METHODS:PBMCsfromhealthypersonswereisolatedandstimulatedbyPMAplusIonomycinatdifferentpe

2、riodsoftime,BrdUwasthenaddedtothecellsonehourbeforetheendofculture.ThecellswereharvestedandstainedwithantiBrdU,anticellsurfaceandintracellularantibodies.Thenthecellswerewashedandanalyzedbyflowcytometer.RESULTS:ThepeakofBrdUincorporationwasobservedinTcellsaftert

3、heywerestimulatedfor48hoursinvitro,butnofurtherincreaseofthepeakofBrdUincorporationwasfoundafterincubatedforalongerperiodoftime.ThecomparisonmadebetweenBrdUincorporationandcellactivationindicatedCD69expressionreachedthepeakafterstimulatedfor8hourswhereasCD25wasat

4、thepeakafterstimulatedfor24hours.Furthermore,nocorrelationbetweenBrdUincorporationandcytokineproduction14wasobserved.HighfrequencyofIFNγproducingcellswasdetectedafterstimulatedfor8hoursbutnoobviousincreasewasobservedforalongerperiodoftime.WhenPBMCwerestimulatedw

5、ithOKT3plusantiCD28,thepercentageofBrdU+TcellswashigherthanthatstimulatedbyPMAplusIonomycin.Similarly,thepercentageofBrdU+CD8+TcellswashigherthanthatofBrdU+CD4+Tcells.CONCLUSION:ThepercentageofBrdU+cellscanbedetectedbyflowcytometertoevaluatetheproliferationofTcel

6、ls.OnlyafewTcellsproliferateafterpolyclonalstimulationandBrdUincorporationisdependentonstuimulantsandtimeofstimulation.Therefore,BrdUincorporationisnotcorrelatedwithactivationmarkersandcytokineproduction.　　[Keywords]BrdU;proliferation;flowcytometer;cellactivation

7、　　通常,人们都是通过3[H]脱氧胸腺嘧啶苷掺入法或MTT比色法来检测T细胞增殖,这些方法测定步骤较多,或敏感性相对较低,且不能从单个细胞水平评价细胞增殖,具有一定限制性。与这些方法不同,脱氧胸腺嘧啶苷的类似物5′溴脱氧尿苷(Bromodeoxyuridine,BrdU)可在细胞合成DNA时替代胸腺嘧啶,14通过固定和破膜,加入荧光标记的抗BrdU抗体,利用流式细胞术检测BrdU掺入率,从而反映细胞增殖水平[1]。因此,BrdU掺入法在检测细胞增殖时,同时还可以检测表面和胞内分子的表达,从而实现了在单个细胞水平分析细

8、胞增殖。本研究我们系统地比较了不同刺激条件下T细胞的增殖及其与细胞活化分子和细胞因子表达的关系,为基础和临床研究提供科学依据。　　1材料和方法　　1.1材料APC标记的抗CD4抗体(APCCD4)、APCCD8、PECD3、APCIFNγ、APCIL2以及相匹配的对照抗体,鼠抗人CD28单克隆抗体（mA