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96 well round bottom tissue culture plates

Company: Sigma-Aldrich
Catalog#: 3799
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Protocol for Isolation, Stimulation and Functional Profiling of Primary and iPSC-derived Human NK Cells
Author:
Date:
2020-12-05
[Abstract]  

Natural killer (NK) cells are innate immune cells, characterized by their cytotoxic capacity, and chemokine and cytokine secretion upon activation. Human NK cells are identified by CD56 expression. Circulating NK cells can be further subdivided into the CD56bright (~10%) and CD56dim NK cell subsets (~90%). NK cell-like cells can also be derived from human induced pluripotent stem cells (iPSC). To study the chemokine and cytokine secretion profile of the distinct heterogenous NK cell subsets, intracellular flow cytometry staining can be performed. However, this assay is challenging when the starting material is limited. Alternatively, NK cell subsets can be enriched, sorted, stimulated, and functionally profiled by measuring secreted effector molecules in the supernatant by Luminex. Here,

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[摘要]  [摘要]天然杀伤(NK)细胞是先天性免疫细胞,其特征在于其细胞毒性能力以及活化后的趋化因子和细胞因子分泌。人NK细胞通过CD56表达鉴定。循环的NK细胞可进一步细分为CD56亮(约10%)和CD56暗NK细胞亚群(约90%)。NK细胞样细胞也可以源自人诱导的多能干细胞(iPSC)。为了研究不同的异源NK细胞亚群的趋化因子和细胞因子分泌概况,可以进行细胞内流式细胞仪染色。然而,当起始原料有限时,该测定法具有挑战性。或者,可以通过Luminex测量上清液中分泌的效应子分子来富集,分选,刺激和功能性分析NK细胞亚群。在这里,我们提供了一种快速直接的方案,用于分离和刺激原代NK细胞或iPSC衍生的NK细胞样细胞,并随后检测分泌的细胞因子和趋化因子,这也适用于少量细胞。


[背景]自然杀伤(NK)细胞是先天免疫系统的一部分,提供第一线防御病毒感染和畸形。在人血中,可以基于CD56和CD16表达鉴定出两个不同的NK细胞群体:CD56明亮的CD16 +/-和CD56暗的CD16 + ...

In vitro Homeostatic Proliferation of Human CD8 T Cells
Author:
Date:
2017-11-20
[Abstract]  Long-lived T-cell–mediated immunity requires persistence of memory T cells in an antigen-free environment while also maintaining a heightened capacity to recall effector functions. Such antigen-independent homeostatic proliferation is mediated in part by the common gamma-chain cytokines IL-7 and IL-15. To further explore the mechanisms governing maintenance of effector functions in long-lived memory T cells during antigen-independent proliferation, human naïve and memory CD8 T cells can be sorted from peripheral blood mononuclear cells (PBMCs), labeled with the proliferation-tracking dye carboxyfluorescein succinimidyl ester (CFSE), and then purified based on their levels of cell division. This allows investigators to assess differences in the desired molecular target in cells that have ... [摘要]  长寿T细胞介导的免疫需要在无抗原环境中持续存在记忆T细胞,同时还保持提高的回忆效应器功能的能力。这种不依赖于抗原的体内平衡增殖部分由常见的γ链细胞因子IL-7和IL-15介导。为了进一步探索在抗原非依赖性增殖过程中维持长效记忆T细胞中效应子功能的机制,可以从外周血单核细胞(PBMCs)中分选人幼稚和记忆CD8T细胞,用增殖跟踪染料羧基荧光素琥珀酰亚胺酯(CFSE),然后根据它们的细胞分裂水平进行纯化。这使得研究人员能够评估经历细胞因子驱动的增殖的细胞中期望的分子靶点的差异。我们在这里提供了一个协议,用于评估在分裂和未分裂的人类幼稚和记忆CD8 T细胞中的表观遗传程序,在用IL-7和IL-15培养7天之后,以说明这种方法如何阐明控制在长寿记忆CD8T细胞的稳态期间保持效应子功能。
【背景】获得性免疫的一个主要特征是对先前遇到的病原体的免疫记忆的发展(Plotkin等人,2013)。记忆CD8 T细胞在为宿主以前遇到的病原体提供终生保护方面发挥重要作用,但为了提供长寿命的保护,T细胞必须已经获得了在无抗原中持续并保持效应功能的能力环境。在此期间,记忆T细胞响应于IL-7/15细胞因子进行抗原非依赖性增殖,尽管程度不同。

记忆CD8 T细胞的总池是几种不同的细胞亚群的异质组合,其对稳态细胞因子的反应不同。例如,CD8 ...

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