| Isolation and Culture of Mouse Lung ILC2s
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Author:
Date:
2018-10-05
[Abstract] Group 2 Innate Lymphoid Cells (ILC2) play an important role in immune responses at barrier surfaces, notably in the lung during airway allergic inflammation or asthma. Several studies have described methods to isolate ILC2s from wild-type naive mice, most of them using cell sorting to obtain a pure population. Here, we describe in detail, a simple, efficient method for isolation and culture of lung mouse ILC2s. Lungs from Rag2-/- mice pretreated with IL-33 are collected and processed into single cell suspensions. Lymphoid cells are then recovered by density gradient separation. Lin-CD45+ cells are selected by depletion of lineage positive cells followed by positive selection of CD45+ cells. Culture of the isolated cells for several days ...
[摘要] 第2组先天性淋巴细胞（ILC2）在屏障表面，特别是在气道过敏性炎症或哮喘期间的肺中的免疫应答中起重要作用。一些研究已经描述了从野生型幼稚小鼠中分离ILC2的方法，其中大多数使用细胞分选来获得纯种群。在这里，我们详细描述了一种简单有效的肺小鼠ILC2分离和培养方法。收集用IL-33预处理的 Rag2 - / - 小鼠的肺并加工成单细胞悬浮液。然后通过密度梯度分离回收淋巴样细胞。通过耗尽谱系阳性细胞然后阳性选择CD45 + 细胞来选择Lin - CD45 + 细胞。将分离的细胞培养数天导致高度纯化的ILC2群体表达典型的细胞表面标志物（CD90.2，Sca1，CD25，CD127和IL-33R）。这些细胞可在培养物中扩增长达10天，并用于多种离体测定或体内过继转移实验。
【背景】第2组先天性淋巴细胞（ILC2）是组织驻留细胞，其在抗寄生虫先天免疫以及过敏性炎症的发展中起关键作用。它们通过产生大量的2型细胞因子IL-5和IL-13对上皮细胞衍生的细胞因子如白细胞介素-33（IL-33）起反应，后者又诱导嗜酸性粒细胞增多和粘液产生（Cayrol和Girard，2018）。为了更好地表征这些细胞的功能和调节，许多组通过荧光激活细胞分选（FACS）从野生型小鼠（WT）的肺中分选ILC2。由于稳定状态下肺中存在的ILC2数量较少，因此该方法导致纯化细胞的产量较低（每只小鼠1×10 ...
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| Ex vivo Natural Killer Cell Cytotoxicity Assay
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Author:
Date:
2013-08-20
[Abstract] Natural Killer (NK) cells are cytotoxic lymphocytes that constitute a major component of the innate immune system. Immunosurveillance of the host by NK cells for malignant and virally-infected cells results in direct cytotoxicity and the production of cytokines to enhance the immune response. This protocol will describe the “gold standard” chromium release assay for measuring the target cell killing capacity of NK cells. Key features of this cytotoxicity assay are that it is performed with sorted NK cells as the effectors and any Major Histocompatibility Class I (MHC-I)-low or deficient tumor cell line can be used as the target cells.
[摘要] 天然杀伤（NK）细胞是细胞毒性淋巴细胞，其构成先天性免疫系统的主要组分。 通过NK细胞对恶性和病毒感染的细胞的宿主的免疫监视导致直接的细胞毒性和细胞因子的产生，以增强免疫应答。 该方案将描述用于测量NK细胞的靶细胞杀伤能力的"金标准"铬释放测定。 该细胞毒性测定的主要特征是其使用分选的NK细胞作为效应物进行，任何主要组织相容性I类（MHC-1） - 低或缺陷的肿瘤细胞系可以用作靶细胞。
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| Natural Killer Cell Transfer Assay
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Author:
Date:
2013-08-20
[Abstract] Natural Killer (NK) cells are cytotoxic lymphocytes that constitute a major component of the innate immune system. Immunosurveillance of the host by NK cells for malignant and virally-infected cells results in direct cytotoxicity and the production of cytokines to enhance the immune response. This protocol will describe the adoptive transfer of purified NK cells into NK cell-deficient tumor bearing mice in order to establish the intrinsic functionality of NK cells.
[摘要] 天然杀伤（NK）细胞是细胞毒性淋巴细胞，其构成先天性免疫系统的主要组分。 通过NK细胞对恶性和病毒感染的细胞的宿主的免疫监视导致直接的细胞毒性和细胞因子的产生，以增强免疫应答。 该方案将描述纯化的NK细胞过继转移到NK细胞缺陷的荷瘤小鼠中以建立NK细胞的内在功能。
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