Abstract：
Microglia are brain-resident phagocytic cells, considered to be the innate immune cells of the central nervous system. Microglia respond to both infectious pathogens in the brain and sterile cellular debris, including the proteinaceous aggregates that accumulate in the brains of patients with Alzheimer\'s disease (AD). Microtubule-associated protein Tau is an intracellular protein that self-aggregates into neurofibrillary tangles in Alzheimer\'s disease and many other neurodegenerative diseases. Ongoing clinical trials are testing whether therapeutic antibodies specific to Tau protein aggregates can reduce pathological protein deposition and improve the course of disease. Data suggest that Tau-specific antibodies act on extracellular Tau aggregates by promoting uptake into microglia cells and thus preventing its prion-like spread to unaffected neurons. Here we describe a protocol to test the effect of Tau-specific antibodies on Tau uptake into microglia by flow cytometry. Recombinant Tau protein is fibrillized in vitro and tagged with a fluorescent label. Then, fibrillized Tau is incubated with the antibody of interest and applied to microglial cells in culture. Uptake of Tau into microglia is then assessed by measuring fluorescence intensity by flow cytometry. With slight modifications, this assay can be used to test effects of many antibodies, various Tau protein compositions, and different microglial sources in a high-throughput format.
摘要：
小胶质细胞是大脑驻留的吞噬细胞，被认为是中枢神经系统的先天免疫细胞。小胶质细胞对大脑中的感染性病原体和无菌细胞碎片都有反应，包括在阿尔茨海默病（AD）患者大脑中积累的蛋白质聚集体。微管相关蛋白Tau是一种在阿尔茨海默病和许多其他神经退行性疾病中自我聚集成神经原纤维缠结的细胞内蛋白。正在进行的临床试验正在测试针对Tau蛋白聚集体的治疗性抗体是否可以减少病理蛋白沉积并改善疾病进程。数据表明，Tau特异性抗体通过促进小胶质细胞的摄取而作用于细胞外Tau聚集体，从而阻止其朊病毒样扩散到未受影响的神经元。在这里，我们描述了一种通过流式细胞术测试Tau特异性抗体对小胶质细胞摄取的影响的方案。重组Tau蛋白在体外原纤维化并用荧光标记标记。然后，将原纤维化的Tau与感兴趣的抗体一起孵育并应用于培养物中的小胶质细胞。然后通过流式细胞术测量荧光强度来评估Tau对小胶质细胞的摄取。稍作修改,这种检测方法可用于检测多种抗体的作用,各种Tau蛋白成分，和高通量形式的不同小胶质细胞来源。
