两种方法分离小鼠脾原始T淋巴细胞效果比较  

Comparison of isolation of primary spleen cells in mice by using Ficoll-Paque gradient cntrifugation and immunomagnetic bead

作　　者：陈嵘祎[1] 阿彩岭[1] 周英[1] 樊翌明[1] 

机构地区：[1]广东医学院附属医院皮肤科,广东湛江524001

出　　处：《中国热带医学》2013年第5期540-542,580,共4页China Tropical Medicine

基　　金：国家自然科学基金面上项目资助(No.81171512)

摘　　要：目的比较Ficoll-Paque梯度离心法与免疫磁珠法分离T淋巴细胞的效果差异,筛选有效的、优异的分选小鼠脾原始T淋巴细胞方法。方法分别采用Ficoll-Paque梯度离心法及Miltenyi Biotec免疫磁珠进行小鼠淋巴细胞分离。血小板计数器统计单核淋巴细胞数,苔盼蓝染色判断细胞存活率,流式细胞术鉴定CD4+CD62+T淋巴细胞纯度,并进行统计学分析。结果免疫磁珠法CD4+CD62+T淋巴细胞纯度较高为(66.27±3.52)%,Ficoll-Paque梯度离心法细胞纯度为(15.27±4.16)%,两者比较差异有统计学意义(P<0.01);但Ficoll-Paque梯度离心法获取淋巴细胞数(1.47±0.35)×108较免疫磁珠法的(0.67±0.26)×108多,两者比较差异有统计学意义(t=8.199,P<0.01),且Ficoll-Paque梯度离心法细胞存活率(93.9±2.6)%较免疫磁珠法的(89.9±2.9)%高,两者比较差异亦有统计学意义(P<0.05)。结论免疫磁珠法适合分离小鼠脾原始T淋巴细胞并用于细胞纯度要求高的免疫学实验,但细胞得率需进一步优化。Objective To compare the effect of isolation of T-lymphocyte of mice by Ficoll-Paque gradient centrifugation and immunomagnetic bead method.Methods Ficoll-Paque gradient centrifugation and Miltenyi Biotec immune magnetic beads were used for isolation of T lymphocyte in mice.Cell number were calculated by platelet counter,cell survival rate was judged by trypan blue dyeing,CD4＋CD62L＋T lymphocyte purity was detected by flow cytometry.Results The purity of CD4＋CD62L＋T lymphocyte in the immune magnetic bead method group was higher（66.27±3.52%）,and lower in Ficoll-Paque gradient centrifugation group（15.27 ±4.16%）,showing significant difference（P〈0.01）,but lymphocyte count in Ficoll-Paque gradient centrifugation group（1.47±0.35×10^8）was more than immune magnetic bead goup（0.67±0.26×10^8）showing significant difference（t=8.199,P〈0.001）.In addition,the cell survival rate in Ficoll-Paque gradient centrifugation group（93.9 ±2.6%）was higher than immunomagnetic bead group（89.9 ±2.9%）also showing significant difference（P〈0.05）.Conclusions Immunomagnetic bead method is suitable for cell purity demanding immunological experiment on condition that further optimization of the cell yield.

关 键 词：鼠 免疫磁珠 淋巴细胞分离 梯度离心 

分 类 号：R332[医药卫生—人体生理学]