| Analysis of Phagosomal Antigen Degradation by Flow Organellocytometry
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Author:
Date:
2016-11-20
[Abstract] Professional phagocytes internalize self and non-self particles by phagocytosis to initiate innate immune responses. After internalization, the formed phagosome matures through fusion and fission events with endosomes and lysosomes to obtain a more acidic, oxidative and hydrolytic environment for the degradation of its cargo. Interestingly, phagosome maturation kinetics differ between cell types and cell activation states. This protocol allows to quantify phagosome maturation kinetics on a single organelle level in different types of phagocytes using flow cytometry. Here, ovalbumin ...
[摘要] 专业吞噬细胞通过吞噬作用内化自身和非自身颗粒以启动先天免疫应答。内化后，形成的吞噬体通过与内体和溶酶体的融合和裂变事件成熟，以获得更酸性，氧化和水解的环境用于其货物的降解。有趣的是，吞噬体成熟动力学在细胞类型和细胞活化状态之间不同。该协议允许使用流式细胞术定量不同类型的吞噬细胞中单个细胞器水平上的吞噬体成熟动力学。在这里，卵白蛋白（OVA）耦合的颗粒用作吞噬作用模型系统在树突状细胞（DC），其通过吞噬内化。在不同的时间点之后，吞噬体成熟参数，例如OVA的吞噬体降解和溶酶体蛋白（例如LAMP-1）的获得，可以通过流式细胞器细胞计数以高度定量的方式同时测量。这些读出可以与其他吞噬体功能相关，例如抗原降解，在DC中的加工和负载。
[背景] ...
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| Vaccine-induced Cytokine Production Detected by Luminex Multiplex Analysis
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Author:
Date:
2014-08-20
[Abstract] Different vaccine and adjuvant combinations are known to rapidly induce antigen presenting cell (APC) maturation and pro-inflammatory cytokine and production, which in turn play an important role in the priming of antigen-specific T cells. Measuring cytokine production systemically in the serum fails to detect localized responses in the lymph nodes draining a subcutaneous immunization site. On the other hand, stimulating APC with vaccine formulations in vitro lacks the complexity of the lymph node microenvironment and the presence of other in vivo factors. Here we analyse ...
[摘要] 已知不同的疫苗和佐剂组合快速诱导抗原呈递细胞（APC）成熟和促炎细胞因子和产生，其反过来在抗原特异性T细胞的引发中起重要作用。 在血清中全身测量细胞因子产生不能检测排出皮下免疫位点的淋巴结中的局部反应。 另一方面，用疫苗制剂在体外刺激APC缺少淋巴结微环境的复杂性和其他体内因子的存在。 在这里我们分析细胞因子生产直接在疫苗引流淋巴结（dLN）提前早期在体内接种。 为此，我们在短暂的离体孵育后对来自整个dLN细胞悬浮液的上清液进行细胞因子多重分析。
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