四聚体技术检测人外周血巨细胞病毒抗原特异性CTL

为深入了解抗病毒免疫机制,探索人在健康状态及巨细胞病毒(CMV)急性感染时抗原特异性细胞毒T淋巴细胞(CTL)数量的动态变化。以CMV抗原肽、HLA-A*0201重链和轻链制备CMV四聚体;分离并检测12名健康被检者外周血单个核细胞(PBMC)中CMV抗原特异CTL数量;PBMC体外培养建立CTL细胞系,于末次刺激的不同时相进行四聚体染色,流式细胞仪(FACS)检测抗原特异CTL数量。结果发现9名被检者PBMC中均检出抗原特异CTL;细胞系中CMV特异性CTL数量急剧增多,细胞系与PBMC相比,差异有显著的统计学意义(P<0.001);研究结果提示,9名被检者感染过CMV,血液中存在少量免疫记忆性T细胞,当再次遭遇同一抗原后发生克隆扩增。

Detection of the cytomegalovirus antigen-specific cytotoxic T lymphocytes in human peripheral blood mononuclear cells by using MHC class Ⅰ tetramer technique

To investigate the dynamic changes in the quantity of antigen-specific cytotoxic T lymphocytes(CTL)in healthy individuals and acute infection with cytomegalovirus(CMV)for a better understanding on the anti-viral immune mechanism, the CMV-tetramer was prepared by CMV antigen peptide together with heavy and light chain of HLA-A*0201.The peripheral blood mononuclear cells(PBMC)were isolated from blood of 12 healthy donors and cultivated ex vitro to set up the antigen- specific CTL lines and the quantity of antigen-specific CTL was determined by FACS using the CMV tetramer.It was found that low but detectable level of the CMV-specific CD8~ T cells could be detected in PBMC of 9 donors by FACS analysis.How- ever,for the CTL line,the quantity of CMV-specific CTL increased obviously with significant difference in comparison with that of PBMC(P<0.001).It is evident that 9 donors had already suffered from CMV infection with the presence of small a- mount of memory T cells which can specifically respond to reinfections.