Quantification of Proliferating Human Antigen-specific CD4+T Cells using Carboxyfluorescein Succinimidyl Ester
Quantification of Proliferating Human Antigen-specific CD4+T Cells using Carboxyfluorescein Succinimidyl Ester
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DOI：
10.3791/59545
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发表时间：
2019-06-01
影响因子：
1.2
通讯作者：
Mannering, Stuart, I
中科院分区：
文献类型：
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作者：
Di Carluccio, Anthony R.;Tresoldi, Eleonora;Mannering, Stuart, I
本发明描述了一种用于测量人外周血单核细胞（PBMC）中抗原特异性CD4（+）T细胞增殖的基于染料稀释的简单体外方法。稳定、无毒的荧光染料如羧基荧光素琥珀酰亚胺酯（CFSE）的开发允许通过荧光染色的减少将稀有的抗原特异性T细胞与旁观者区分开，如通过流式细胞术检测的。该方法相对于替代方法具有以下优点：（i）它对低频T细胞非常敏感，（ii）不需要抗原或表位的知识，（iii）可以分析应答细胞的表型，以及（iv）可以分选活的应答细胞并用于进一步分析，例如T细胞克隆。
Described is a simple, in vitro, dye dilution-based method for measuring antigen-specific CD4( )(+)T cell proliferation in human peripheral blood mononuclear cells (PBMCs). The development of stable, non-toxic, fluorescent dyes such as carboxyfluorescein succinimidyl ester (CFSE) allows for rare, antigen-specific T cells to be distinguished from bystanders by diminution in fluorescent staining, as detected by flow cytometry. This method has the following advantages over alternative approaches: (i) it is very sensitive to low-frequency T cells, (ii) no knowledge of the antigen or epitope is required, (iii) the phenotype of the responding cells can be analyzed, and (iv) viable, responding cells can be sorted and used for further analysis, such as T cell cloning.