鸡自然杀伤细胞的分离与免疫表型分析  

Isolation and phenotypic analysis of chicken natural killer cells

作　　者：杨超[1] 张晓娜[1] 张园华[2] 廉传江[1] 文辉强[1] 陈洪岩[1] 韩凌霞[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/实验动物与比较医学团队,黑龙江哈尔滨150001 [2]山西农业大学动物科技学院,山西太谷030801

出　　处：《中国预防兽医学报》2014年第3期236-238,共3页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金项目(31101800);科技部支撑项目(2013BAK11B02)

摘　　要：为研究鸡自然杀伤(NK)细胞,本实验采用冷胰酶消化法消化14日胚龄鸡胚脾脏组织,分离收获鸡胚脾脏淋巴细胞,在含有重组鸡白介素-2和条件培养基的IMDM培养液中培养48 h,收获悬浮细胞,暂时命名为TCR0细胞(即鸡NK细胞)。流式前向角和侧向角分析显示培养的TCR0细胞中85.1%的细胞大小均一,颗粒度一致;瑞氏吉姆萨染料染色结果表明TCR0细胞内一侧含有嗜酸性大颗粒,符合鸡NK细胞的形态特征;流式细胞术单克隆抗体染色结果显示,TCR0细胞中,97.4%高表达MHCⅠ类分子,30.8%低表达CD8a,不表达T细胞表面标记CD3和B细胞表面标记CD4和Bu-1,符合鸡NK细胞表面标记特点。本实验是国内成功分离并纯化鸡NK细胞的首次报道。The study of chicken NK （chNK） cells is greatly lagged due to the lack of exclusively expressed markers for chNK cells and the complex methods to isolate and culture for chNK cells. According to the development of chicken immune system, the embryonic spleen in 14 day old-chicken is a rich source of chNK cells. In this experiment, chicken embryonic spleens were digested with cold trypsin to isolate the spleen lymphocytes and then cultured in IMDM medium containing recombinant chicken IL-2 for 48 hours. The harvested suspension cells, provisionally named TCR0 cells （chNK cells）, were subjected to the flow cytometry assay based forward scatter and side scatter which indicated that 85.1% of them were morphologically similar. In addition, the Wright-Giemas’ staining assay also showed that there were large eosinophilic granules in one side of the cells, which is characterized by chNK cells. Furthermore, the immunophenotypic analysis indicated that the harvested chNK cells were unable to express T cell and B cell markers of CD3, CD4 and Bu-1, whereas, the 97.4% of TCR0 cells were stained brightly for MHC class I molecules and 30.8% dimly for CD8a. These results demonstrated that we haven successfully established the protocol to isolate and culture chNK cells.

关 键 词：鸡自然杀伤细胞 分离与培养 免疫表型 

分 类 号：S852.4[农业科学—基础兽医学]