Mabtech AB, Box 1233, SE-131 28 Nacka Strand, Sweden.
Cells. 2014 Nov 27;3(4):1116-30. doi: 10.3390/cells3041116.
The involvement of T-helper (Th)1, Th17 and Th22 cell subsets, in immunity, as well as in pathological inflammatory reactions, makes it important to determine their relative proportion. A triple FluoroSpot detecting the hallmark cytokines of Th1 (IFN-γ), Th17 (IL-17A) and Th22 (IL-22) was developed and evaluated using human peripheral blood mononuclear cells from healthy donors incubated with tetanus toxoid, Candida albicans extract, mycobacterial purified protein derivative or medium only. Antigen stimulation yielded mainly cells secreting IFN-γ, IL-17A or IL-22 alone but lower proportions of double-secreting cells were also found; triple-secreting cells were rare. The response to C. albicans contrasted in that higher proportions of IL-17A single secreting as well as co-secreting cells, in particular IL-17A/IL-22, were found. The FluoroSpot analysis correlated well with single cytokine ELISpot assays ran in parallel and the methods displayed a comparable sensitivity. The results demonstrate the functionality of the FluoroSpot assay for simultaneous analysis of distinct Th1, Th17, Th22 as well as intermediate cell populations. The method provides a mean for a simple and rapid analysis of the involvement of these cells in immunity and disease.
辅助性 T 细胞（Th）1、Th17 和 Th22 亚群的参与，不仅涉及免疫，还涉及病理性炎症反应，因此确定它们的相对比例很重要。本研究开发并评估了一种三重 FluoroSpot 检测方法，该方法可检测 Th1（IFN-γ）、Th17（IL-17A）和 Th22（IL-22）的标志性细胞因子，其检测对象为人外周血单个核细胞，这些细胞来自于破伤风类毒素、白色念珠菌提取物、分枝杆菌纯蛋白衍生物或仅用培养基孵育的健康供体。抗原刺激主要产生单独分泌 IFN-γ、IL-17A 或 IL-22 的细胞，但也发现了比例较低的双分泌细胞；三重分泌细胞很少见。对白念珠菌的反应则相反，发现了更高比例的 IL-17A 单独分泌以及共分泌细胞，特别是 IL-17A/IL-22。FluoroSpot 分析与平行运行的单个细胞因子 ELISA 检测结果相关性良好，并且两种方法的灵敏度相当。结果表明，FluoroSpot 分析可用于同时分析不同的 Th1、Th17、Th22 以及中间细胞群，该方法为分析这些细胞在免疫和疾病中的参与提供了一种简单快速的手段。
