以LRS滤器获取白细胞作为研究用T细胞的可行性分析  

Replacing human peripheral blood leukocytes with LRS-separated leukocytes for scientific research

作　　者：古彦铮[1,2] 汤龙海[1] 黄子逸[2] 李维洁[1] 张光波[2] 王明元[1] 

机构地区：[1]苏州市中心血站,215006 [2]苏州大学附属第一医院临床免疫研究所,215006

出　　处：《中华微生物学和免疫学杂志》2016年第8期607-614,共8页Chinese Journal of Microbiology and Immunology

基　　金：苏州市科技计划项目（SYS210424）;国家自然科学基金青年项目（81402547）

摘　　要：目的：分析单采血小板去白细胞（ leukoreduction system, LRS）滤器收集的白细胞替代常规全血人外周血白细胞的可行性,并对两个来源的T细胞进行了体外生物学和表型比较分析。方法本实验用同一志愿者LRS滤器中富集的白细胞和血液检测后剩余的全血经淋巴细胞分离液（ Ficoll）分离成单个核细胞,用流式细胞仪对细胞表型比较分析。同时经磁珠分选出CD3^＋T细胞,加入CD3、CD28激发型抗体与IL-2体外扩增培养10 d,并以细胞计数、流式细胞仪检测以及CBA细胞因子分泌测定等方法,对比LRS滤器血与全血分离出T细胞表型和生物学功能的差异。结果 LRS滤器血与全血分离出的细胞,初始状态表型相似度极高；LRS滤器血的CD3^＋T细胞分选率非常高,达到了实验要求。经扩增后的细胞的数量、表型、共刺激分子的表达、细胞因子分泌的数值都很相似,差异无统计学意义。结论本研究证实了LRS滤器血分离的T细胞在活力、表型和功能方面可以替代全血中的T细胞应用于基础实验研究,为解决临床用血和实验用血的矛盾提供了新途径。Objective To investigate the feasibility of using leukocytes that were filtered out by LeukoReduction System （ LRS） to replace conventional human peripheral blood leukocytes in experimental researches and to comparatively analyze the differences between them in vitro biological functions and pheno-types of T cells. Methods Mononuclear cells were isolated from LRS-separated leukocytes and whole blood sample that collected from the same person by using Ficoll. Fluorescence-activated cell sorting （ FACS） was performed to analyze the phenotypes of T cells. CD3^＋T cells were sorted out by using magnetic beads. The T cells that were collected by using two different ways were incubated with anti-CD3 and anti-CD28 antibodies and IL-2 in vitro for 10 days. Several assays including cell counting, FACS and cytometric beads array （ CBA） were performed to comparatively analyze the differences in biological functions and phenotypes of T cells that were isolated by different methods. Results The phenotypes of T cells isolated from LRS filter and whole blood sample were highly similar at the initial stage. The sorting rate of CD3^＋T cells form LRS filter reached a high level and met the requirements for experimental researches. No statistically significant differ-ences in cell count, phenotype, expression of costimulatory molecules and cytokine secretion were observed between T cells isolated from LRS filter and whole blood sample. Conclusion This study suggested that the T cells isolated from LRS filter could be used as an alternative to whole blood T cells for fundamental resear-ches since they were similar in cell vitality, phenotype and biological functions. It provided a new way to solve the problem of blood shortage in clinic and scientific research.

关 键 词：单采血小板去白细胞滤器 外周血 CD3^+ T细胞 

分 类 号：R392[医药卫生—免疫学]