流式细胞仪实时监测细胞内Ca^(2+)浓度变化  

Real-time monitoring of intracellular Ca^(2+) concentration by flow cytometry

作　　者：曹云新[1] 李豫容 杨安钢[1] 

机构地区：[1]解放军第四军医大学免疫学教研室,陕西省西安市710032 [2]解放军第四军医大学预防医学系放射医学教研室,陕西省西安市710032

出　　处：《中国组织工程研究与临床康复》2010年第24期4481-4483,共3页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家自然科学基金(30700730)资助~~

摘　　要：背景:多种信号转导途径可影响细胞内Ca2+浓度的变化,因而对细胞内钙离子变化的检测,有助于了解细胞功能的启动、加强或抑制。目的:利用流式细胞术实时监测细胞内钙离子浓度的动态变化。方法:选取健康人新鲜全血,用淋巴细胞分离液分离淋巴细胞,使用Ca2+荧光指示剂Fluo-3/AM负载淋巴细胞,加入CD3单克隆抗体和CD28单克隆抗体刺激淋巴细胞活化,以流式细胞仪动态监测活化淋巴细胞内Ca2+的浓度变化。结果与结论:T淋巴细胞在静息状态时,钙离子的荧光值平稳,显示为基线;加入抗CD3单克隆抗体和抗CD28单克隆抗体后,T淋巴细胞被活化,钙离子的浓度迅速升高,持续大约两三分钟后下降,逐渐趋向平缓。结果提示采用钙离子指示剂Fluo-3负载细胞与流式细胞术分析可以跟踪细胞内钙离子的动态变化,成为钙离子动态变化监测的有效方法。BACKGROUND：The concentration changes of Ca2＋ in cells is susceptible to various signal transduction pathways,thus,it is helpful to monitor changes in Ca2＋ for a better understanding of the priming,augmentation or inhibition of the functions of T-cells.OBJECTIVE：To detect dynamic changes in Ca2＋ using flow cytometry.METHODS：The fresh whole blood was obtained from human,and the lymphocytes were separated.CD3mAb and CD28mAb were added into the Fluo-3/AM-loaded cells,and the changes of Ca2＋ concentration were real-time monitored by flow cytometry.RESULTS AND CONCLUSION：The fluorescence value of Ca2＋ was stable and showed a baseline when the T lymphocytes at static status;after adding anti-CD3mAb and anti-CD28mAb,the T lymphocytes were activated,and the Ca2＋ concentration was quickly increased,lasted for 2-3 minutes,and gradually trended to be stale.The results revealed that flow cytometric analysis enabled us to trace dynamic movement of Ca2＋ using Fluo-3,which can be used as an effective method for monitoring Ca2＋ changes.

关 键 词：细胞内CA2+ 浓度 流式细胞仪 实时监测 淋巴细胞 

分 类 号：R318[医药卫生—生物医学工程]