Multicolor flow cytometry analyses of cellular immune response in rhesus macaques.
Multicolor flow cytometry analyses of cellular immune response in rhesus macaques.
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DOI：
10.3791/1743
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发表时间：
2010-04-22
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影响因子：
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通讯作者：
Sastry, K Jagannadha
中科院分区：
文献类型：
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作者：
He, Hong;Courtney, Amy N;Sastry, K Jagannadha
恒河猴模型是目前在了解发病机制以及开发疫苗和治疗方法方面最好的HIV-AIDS模型（1，2，3）。在这里，我们描述了一种方法，用于细胞免疫反应的详细表型和功能分析，特别是CD 4+和CD 8 + T细胞以及个体记忆亚群的细胞内细胞因子产生。我们获得了精确的定量和定性测量干扰素γ（INF-）和白细胞介素（IL）-2的生产在CD 4+和CD 8 + T细胞从恒河猴PBMC与PMA加离子霉素（PMA+I）刺激。细胞因子谱在不同的记忆细胞亚群中是不同的。此外，该方案为我们提供了灵敏度，以证明来自用我们实验室开发的HIV包膜肽混合物疫苗免疫的恒河猴的PBMC样品中抗原特异性CD 4+和CD 8 + T细胞亚群的即使很小部分。流式细胞术技术是一种强有力的工具，可以在非特异性或抗原特异性刺激（5，7）后精确鉴定具有产生精氨酸能力（6）的不同T细胞群（4，5）。
The rhesus macaque model is currently the best available model for HIV-AIDS with respect to understanding the pathogenesis as well as for the development of vaccines and therapeutics(1,2,3). Here, we describe a method for the detailed phenotypic and functional analyses of cellular immune responses, specifically intracellular cytokine production by CD4+ and CD8+ T cells as well as the individual memory subsets. We obtained precise quantitative and qualitative measures for the production of interferon gamma (INF-) and interleukin (IL) -2 in both CD4+ and CD8+ T cells from the rhesus macaque PBMC stimulated with PMA plus ionomycin (PMA+I). The cytokine profiles were different in the different subsets of memory cells. Furthermore, this protocol provided us the sensitivity to demonstrate even minor fractions of antigen specific CD4+ and CD8+ T cell subsets within the PBMC samples from rhesus macaques immunized with an HIV envelope peptide cocktail vaccine developed in our laboratory. The multicolor flow cytometry technique is a powerful tool to precisely identify different populations of T cells (4,5) with cytokine-producing capability(6) following non-specific or antigen-specific stimulation (5,7).