A novel method for isolation of human lung T cells from lung resection tissue reveals increased expression of GAPDH and CXCR6
A novel method for isolation of human lung T cells from lung resection tissue reveals increased expression of GAPDH and CXCR6
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DOI：
10.1016/j.jim.2008.12.001
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发表时间：
2009-03-15
影响因子：
2.2
通讯作者：
Guillen, C.
中科院分区：
文献类型：
--
作者：
Day, C. E.;Zhang, S. D.;Guillen, C.
肺 T 淋巴细胞在肺部免疫和炎症中很重要。由于受到其他细胞类型（主要是肺泡巨噬细胞）的污染，研究这些细胞一直很困难。我们开发了一种结合多种方法从肺切除组织中分离肺 T 细胞的新方法。首先，将肺组织切碎并通过尼龙网过滤。通过 Percoll 密度离心富集淋巴细胞，并使用人 CD3 微珠纯化 T 细胞，产生 90.5% +/- 1.9% (n = 11) 的纯淋巴细胞。粗细胞制剂的 T 细胞产量为 10.8 +/- 2.1%，使用碘化丙啶 (PI) 染色和台盼蓝计算的活力通常超过 95%。纯化过程不影响 CD69 或 CD103 的表达，起始群体和纯化细胞之间的 CD4 和 CD8 细胞比例也不存在差异。微阵列分析和实时 RT-PCR 显示，与血液来源的 T 细胞相比，肺 T 细胞的 GAPDH 和 CXCR6 上调。该技术高度富集肺 T 细胞，以便对这些细胞的生物学进行详细研究。 (C) 2008 Elsevier B.V. 保留所有权利。
Lung T lymphocytes are important in pulmonary immunity and inflammation. it has been difficult to study these cells due to contamination with other cell types, mainly alveolar macrophages. We have developed a novel method for isolating lung T cells from lung resection tissue, using a combination of approaches. Firstly the lung tissue was finely chopped and filtered through a nylon mesh. Lymphocytic cells were enriched by Percoll density centrifugation and the T cells purified using human CD3 microbeads, resulting in 90.5% +/- 1.9% (n = 11) pure lymphocytes. The T cell yield from the crude cell preparation was 10.8 +/- 2.1% and viability, calculated using propidium iodide (PI) staining and trypan blue, was typically over 95%. The purification process did not affect expression of CD69 or CD103, nor was there a difference in the proportion of CD4 and CD8 cells between the starting population and the purified cells. Microarray analysis and real time RT-PCR revealed upregulation of GAPDH and CXCR6 of the lung T cells as compared to blood-derived T cells. This technique highly enriches lung T cells to allow detailed investigation of the biology of these cells. (C) 2008 Elsevier B.V. All rights reserved.