Isolation and Activation of Murine Lymphocytes
Isolation and Activation of Murine Lymphocytes
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DOI：
10.3791/54596
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发表时间：
2016-10-01
影响因子：
1.2
通讯作者：
Su, I-hsin
中科院分区：
文献类型：
--
作者：
Lim, Jun Feng;Berger, Heidi;Su, I-hsin
B 细胞和 T 细胞具有极其多样化的抗原受体库，能够针对几乎所有入侵病原体发起特异性免疫反应 (1,2)。可以理解的是，这种复杂的能力是由参与各种细胞过程的大量分子控制的，以确保及时和空间调节的免疫反应(3)。在这里，我们描述了使用磁性细胞分选技术快速分离高度纯化的小鼠淋巴细胞的实验程序。然后可以对所得纯化淋巴细胞进行各种体外或体内功能测定，例如通过免疫印迹测定刺激后淋巴细胞信号传导能力(4)，以及通过 H-3-胸苷掺入或羧基荧光素二乙酸琥珀酰亚胺酯 (CFSE) 标记研究增殖能力 (5-7)。除了比较对照淋巴细胞和转基因淋巴细胞的功能能力外，我们还可以确定体内抗原呈递细胞 (APC) 的 T 细胞刺激能力，如我们使用移植的 CFSE 标记的 OT-I T 细胞的代表性结果所示。
B and T cells, with their extremely diverse antigen-receptor repertoires, have the ability to mount specific immune responses against almost any invading pathogen(1,2). Understandably,such intricate abilities are controlled by a large number of molecules involved in various cellular processes to ensure timely and spatially regulated immune responses(3). Here, we describe experimental procedures that allow rapid isolation of highly purified murine lymphocytes using magnetic cell sorting technology. The resulting purified lymphocytes can then be subjected to various in vitro or in vivo functional assays, such as the determination of lymphocyte signaling capacity upon stimulation by immunoblotting(4) and the investigation of proliferative abilities by H-3-thymidine incorporation or carboxyfluorescein diacetate succinimidyl ester (CFSE) labeling(5-7). In addition to comparing the functional capacities of control and genetically modified lymphocytes, we can also determine the T cell stimulatory capacity of antigen-presenting cells (APCs) in vivo, as shown in our representative results using transplanted CFSE-labeled OT-I T cells.