Optimized lymphocyte isolation methods for analysis of chemokine receptor expression
Optimized lymphocyte isolation methods for analysis of chemokine receptor expression
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DOI：
10.1016/s0022-1759(03)00186-8
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发表时间：
2003-08-01
影响因子：
2.2
通讯作者：
Roederer, M
中科院分区：
文献类型：
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作者：
Berhanu, D;Mortari, F;Roederer, M
通常用于从外周血中分离富集的淋巴细胞群的操作可影响趋化因子受体的测量水平。因此，受体表达细胞亚群的最佳灵敏度和准确区分需要对表达水平影响最小的细胞分离方法。我们使用流式细胞术检测不同的方案处理和染色T淋巴细胞对趋化因子受体表达的影响。我们的研究结果限制了一些趋化因子受体的FACS分析在标准方法（如Ficoll密度分离）后会受到影响。虽然最佳方法通常是在裂解全血之前对细胞进行染色，但这在许多实验条件下可能不实用。总的来说，我们发现在Ficoll分离后在37 ℃下染色细胞产生了优异的结果。然而，使用的精确方法将取决于测量的受体。我们使用最佳的方法来比较幼稚和记忆T细胞亚群上的趋化因子受体的表达，使用8色流式细胞术。由爱思唯尔公司出版
Manipulations typically used to isolate enriched lymphocyte populations from peripheral blood can impact on the measured levels of chemokine receptors. Optimum sensitivity and accurate discrimination of receptor-expressing cell subsets therefore requires cell isolation methods that minimally affect expression levels. We used flow cytometry to examine the effects of different protocols for processing and staining T lymphocytes on chemokine receptor expression. Our results confine that FACS analysis of some chemokine receptors is compromised after standard methods (such as Ficoll density separation). While the optimal method was typically to stain cells prior to lysing whole blood, this may not be practical in many experimental conditions. In general, we found that staining cells at 37C following Ficoll separation yielded excellent results. However, the precise method used will depend on which receptor is being measured. We used the optimal methods to compare the expression of chemokine receptors on naive and memory T-cell subsets using 8-color flow cytometry. Published by Elsevier B.V.