Characterization of immune responses is currently hampered by the lack of systems enabling quantitative and dynamic phenotypic characterization of individual cells and, in particular, analysis of secreted proteins such as cytokines and antibodies. We recently developed a simple and robust microfluidic platform, DropMap, to measure simultaneously the kinetics of secretion and other cellular characteristics, including endocytosis activity, viability and expression of cell-surface markers, from tens of thousands of single immune cells. Single cells are compartmentalized in 50-pL droplets and analyzed using fluorescence microscopy combined with an immunoassay based on fluorescence relocation to paramagnetic nanoparticles aligned to form beadlines in a magnetic field. The protocol typically takes 8–10 h after preparation of microfluidic chips and chambers, which can be done in advance. By contrast, enzyme-linked immunospot (ELISPOT), flow cytometry, time-of-flight mass cytometry (CyTOF), and single-cell sequencing enable only end-point measurements and do not enable direct, quantitative measurement of secreted proteins. We illustrate how this system can be used to profile downregulation of tumor necrosis factor-α (TNF-α) secretion by single monocytes in septic shock patients, to study immune responses by measuring rates of cytokine secretion from single T cells, and to measure affinity of antibodies secreted by single B cells.

目前，由于缺乏能够对单个细胞进行定量和动态表型表征的系统，尤其是对细胞因子和抗体等分泌蛋白的分析，免疫反应的表征受到了阻碍。我们最近开发了一个简单而强大的微流体平台 DropMap，用于同时测量来自数万个免疫细胞的分泌动力学和其他细胞特征，包括细胞内吞活性、活力和细胞表面标记物的表达。单个细胞被分隔成 50 pL 液滴，并使用荧光显微镜结合基于荧光重定位的免疫测定法对顺磁性纳米粒子进行分析，这些纳米粒子在磁场中对齐形成珠线。该协议通常在准备好微流控芯片和腔室后需要 8-10 小时，这可以提前完成。相比之下，酶联免疫斑点 (ELISPOT)、流式细胞术、飞行时间质谱细胞术 (CyTOF) 和单细胞测序只能进行终点测量，不能直接定量测量分泌的蛋白质。我们说明了该系统如何用于分析感染性休克患者中单个单核细胞分泌的肿瘤坏死因子-α (TNF-α) 的下调，通过测量单个 T 细胞的细胞因子分泌率来研究免疫反应，以及测量亲和力由单个 B 细胞分泌的抗体。和单细胞测序只能进行终点测量，不能直接定量测量分泌的蛋白质。我们说明了该系统如何用于分析感染性休克患者中单个单核细胞分泌的肿瘤坏死因子-α (TNF-α) 的下调，通过测量单个 T 细胞的细胞因子分泌率来研究免疫反应，以及测量亲和力由单个 B 细胞分泌的抗体。和单细胞测序只能进行终点测量，不能直接定量测量分泌的蛋白质。我们说明了该系统如何用于分析感染性休克患者中单个单核细胞分泌的肿瘤坏死因子-α (TNF-α) 的下调，通过测量单个 T 细胞的细胞因子分泌率来研究免疫反应，以及测量亲和力由单个 B 细胞分泌的抗体。