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LIVE/DEADTM Fixable Aqua Dead Cell Stain Kit, for 405 nm excitation

LIVE / DEAD ®可固定水性死细胞染色试剂盒

Company: Thermo Fisher Scientific
Catalog#: L34957
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Intravenous Labeling and Analysis of the Content of Thymic Perivascular Spaces
Author:
Date:
2018-03-05
[Abstract]  Following development in the thymus, T cells are thought to exit into the periphery predominantly through perivascular spaces (PVS). This exit route is used by conventional T cells, and likely also applies to unconventional T cell subsets, such as precursors of CD8αα and TCRγδ intraepithelial lymphocytes, regulatory T cells and natural killer T cells. Additional cell types might also be found in the PVS and initiate interactions with exiting T cells. The exact content of the PVS, and the processes within, are not well studied. To distinguish vascular from resident cells within various tissues by flow cytometry, intravenous (i.v.) labeling is becoming a commonly employed method. We recently used anti-CD45.2 antibodies and magnetic enrichment to further evaluate this technique, and compared ... [摘要]  在胸腺发育后,T细胞被认为主要通过血管周围间隙(PVS)进入周边。 这种退路由常规T细胞使用,也可能适用于非常规T细胞亚群,例如CD8αα和TCRγδ上皮内淋巴细胞的前体,调节性T细胞和天然杀伤T细胞。 其他细胞类型也可能在PVS中发现并启动与退出T细胞的相互作用。 PVS的确切内容及其内部过程尚未得到充分研究。 为了通过流式细胞术将血管与各种组织中的驻留细胞区分开,静脉内(静脉内)标记正在成为常用方法。 我们最近使用抗CD45.2抗体和磁性富集来进一步评估这种技术,并比较胸腺和血液中的标记和未标记的细胞。 该测定可用于特异性研究胸腺PVS内的造血细胞亚群。

【背景】未成熟的胸腺细胞经历一系列成熟步骤,包括正向和负向选择,其消除了大部分发育中的T细胞。由此产生的成熟T细胞库因此形成朝向更高比例的有益克隆和减少比例的危险自反应克隆。胸腺还产生较少丰富的T细胞亚群,其通常用于维持免疫系统,组织和代谢稳态,包括:TCRγδ细胞,调节性T细胞(Treg),自然杀伤T细胞(NKT),上皮内淋巴细胞(IEL)和粘膜相关不变T(MAIT)细胞。成熟的胸腺细胞准备迁移到外周,上调表达识别鞘氨醇-1磷酸(S1P)的受体(S1PR1)的表达,S1P是血液中高浓度存在的脂质分子。 S1PR1 + T细胞沿着S1P梯度迁移并卷入血管循环中。

胸腺血管周围间隙(PVS)是实质和脉管系统之间的基膜分隔室。它们被认为促进了细胞的运输,特别是从胸腺移出的成熟T细胞(Mori等人,2007; ...

Mouse CD8+ T Cell Migration in vitro and CXCR3 Internalization Assays
Author:
Date:
2017-03-20
[Abstract]  Chemokines are molecules that regulate the positioning of cells during homeostasis and inflammation. CXCL10 is an interferon-induced chemokine that attracts cells that express the chemokine receptor CXCR3 on their surface. CXCL10 expression is often induced upon inflammation and guides lymphocytes, such as T and NK cells, into the injured tissues. Notably, CXCL10 binding to CXCR3 induces receptor internalization and, therefore, low CXCR3 levels in cells positive for CXCR3 expression can be indicative of chemokine signaling.

Here, we describe an in vitro method to evaluate the ability of murine CD8+ T cells to migrate towards recombinant murine CXCL10; and a flow cytometry assay to measure CXCR3 expression levels at the surface of T cells, after exposure to ...
[摘要]  趋化因子是调节体内平衡和炎症期间细胞定位的分子。 CXCL10是干扰素诱导的趋化因子,其吸引在其表面上表达趋化因子受体CXCR3的细胞。 CXCL10表达通常在炎症诱导并引导淋巴细胞如T和NK细胞进入受损组织。值得注意的是,CXCL10与CXCR3结合诱导受体内化,因此CXCR3表达阳性细胞中的CXCR3水平降低可能是趋化因子信号传导的指示。
 这里,我们描述体外方法来评估鼠CD8 + T细胞向重组鼠CXCL10迁移的能力;以及暴露于不同剂量的趋化因子后在T细胞表面测量CXCR3表达水平的流式细胞术测定。

背景 趋化因子介导的T细胞运输是稳态和炎症期间的重要过程。活化的CD8 + T细胞表达趋化因子受体,例如CXCR3,允许它们向趋化因子CXCL9,10和11迁移,通常在损伤组织上上调。调节T细胞迁移的分子线索的评估对于了解其功能背后的生物学非常重要,但是在体内运行的复杂机制有时难以去卷积。在这里,我们提供有关体外方法的详细信息,以评估CD8 + T细胞上的趋化因子功能,重点是CXCL10介导的化学吸引和CXCR3内化。我们使用可以容易地在体外扩增和活化的抗原特异性转基因CD8 +细胞,因此提供足够数量的表型相同的淋巴细胞(例如, ...

Ex vivo Human Natural Killer (NK) Cell Stimulation and Intracellular IFNγ and CD107a Cytokine Staining
Author:
Date:
2015-06-20
[Abstract]  Natural killer (NK) cells comprise 5–20% of peripheral blood mononuclear cells (PBMC) in humans. In addition to their fundamental roles in the defense against viral infections and tumor surveillance, NK cells help shape adaptive immune responses through their production of cytokines. NK cells are traditionally identified as CD3neg, CD14neg, CD19neg lymphocytes expressing CD56. Using a combination of markers that includes CD56 and CD7 greatly increases the ability to define the phenotype and function of NK cell subsets. Two key markers of NK cell function are the production of IFNγ and the release of cytotoxic granules measured by the expression of CD107a. Here we describe a method to assess IFNγ and CD107a expression in NK cells following stimulation with ... [摘要]  自然杀伤(NK)细胞在人中包含5-20%的外周血单核细胞(PBMC)。 除了它们在防御病毒感染和肿瘤监测中的基本作用,NK细胞通过其细胞因子的产生帮助形成适应性免疫应答。 NK细胞传统上被鉴定为表达CD56的CD3阴性,CD14阳性,CD19阴性淋巴细胞。 使用包括CD56和CD7的标记物的组合极大地增加了定义NK细胞亚群的表型和功能的能力。 NK细胞功能的两个关键标记是IFNγ的产生和通过CD107a的表达测量的细胞毒性颗粒的释放。 在这里我们描述了一种方法来评估在靶细胞或细胞因子刺激后NK细胞中的IFNγ和CD107a表达。 该方法可用于评估来自广泛的研究参与者的外周血单核细胞中NK细胞的一般功能能力。

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