中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (24): 4481-4483.doi: 10.3969/j.issn.1673-8225.2010.24.025

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

流式细胞仪实时监测细胞内Ca 2+浓度变化

曹云新1，李豫容2，杨安钢1   

  1. 解放军第四军医大学，1免疫学教研室，2预防医学系放射医学教研室，陕西省西安市   710032
  • 出版日期:2010-06-11 发布日期:2010-06-11
  • 作者简介:曹云新，女，1959年生，山东省费县人，汉族，1997年解放军第四军医大学毕业，高级实验师，主要从事流式细胞术和细胞分析研究。 Caoxin87@fmmu.edu.cn
  • 基金资助:

    课题受国家自然科学基金(30700730)资助。

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

Cao Yun-xin1, Li Yu-rong2, Yang An-gang1   

  1. 1 Department of Immunology, 2 Department of Preventive Medicine, the Fourth Military Medical University, Xi’an  710032, Shaanxi Province, China
  • Online:2010-06-11 Published:2010-06-11
  • About author:Cao Yun-xin, Senior Experimentalist, Department of Immunology, the Fourth Military Medical University, Xi’an 710032, Shaanxi Province, China Caoxin87@fmmu.edu.cn
  • Supported by:

    the National Natural Science Foundation of China, No. 30700730*

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摘要：

背景：多种信号转导途径可影响细胞内Ca 2+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Ca 2+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Ca 2+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。

关键词: 细胞内Ca 2+ , 浓度, 流式细胞仪, 实时监测, 淋巴细胞

Abstract:

BACKGROUND: The concentration changes of Ca 2+ in cells is susceptible to various signal transduction pathways, thus, it is helpful to monitor changes in Ca 2+ for a better understanding of the priming, augmentation or inhibition of the functions of T-cells.
OBJECTIVE: To detect dynamic changes in Ca 2+ 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 Ca 2+ concentration were real-time monitored by flow cytometry.
RESULTS AND CONCLUSION: The fluorescence value of Ca 2+ 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 Ca 2+ 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 Ca 2+ using Fluo-3, which can be used as an effective method for monitoring Ca 2+ changes. 

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