| In vitro Assessment of Immunological Synapse Formation by Flow Cytometry
|
|
Author:
Date:
2016-03-20
[Abstract] In adaptive immune system, formation of immunological synapse between T cells and antigen presenting cells (dendritic cells, B cells, and macrophages) or target cells (tumor cells and viral-infected cells) is critical for the execution of T cell immune responses via cytokine secretion or direct killing activity. Here, we describe the practical methods that directly measure the number of conjugates as a result of immunological synapse formation between T cells and superantigen-loaded B cells or between cytotoxic T cells and antigen-loaded target cells by dual-color flow cytometry.
[摘要] 在适应性免疫系统中，T细胞和抗原呈递细胞（树突细胞，B细胞和巨噬细胞）或靶细胞（肿瘤细胞和病毒感染细胞）之间的免疫突触的形成对通过细胞因子执行T细胞免疫应答至关重要 分泌或直接杀死活动。 在这里，我们描述了通过双色流式细胞术直接测量T细胞和加载超抗原的B细胞之间或细胞毒性T细胞和抗原负载的靶细胞之间的免疫突触形成的结合物数量的实际方法。
|
|
|
| Quantification of T Cell Antigen-specific Memory Responses in Rhesus Macaques, Using Cytokine Flow Cytometry (CFC, also Known as ICS and ICCS): from Assay Set-up to Data Acquisition
|
|
Author:
Date:
2014-04-20
[Abstract] What was initially termed ‘CFC’ (Cytokine Flow Cytometry’) is now more commonly known as ‘ICS’ (Intra Cellular Staining), or less commonly as ‘ICCS’ (Intra Cellular Cytokine Staining). The key innovations were use of an effective permeant (allowing intracellular staining), and a reagent to disrupt secretion (trapping cytokines, thereby enabling accumulation of detectable intracellular signal). Because not all researchers who use the technique are interested in cytokines, the ‘ICS’ term has gained favor, though ‘CFC’ will be used here.
CFC is a test of cell function, exposing lymphocytes to antigen in culture, then measuring any cytokine responses elicited. Test cultures are processed so as to stain cells with monoclonal antibodies tagged with fluorescent markers, and to ...
[摘要] 最初称为"CFC"（细胞因子流式细胞术）的现在更通常称为"ICS"（细胞内染色），或更不常称为"ICCS"（细胞内细胞因子染色）。关键的创新是使用有效的渗透物（允许细胞内染色）和试剂破坏分泌（捕获细胞因子，从而能够累积可检测的细胞内信号）。因为不是所有使用该技术的研究人员都对细胞因子感兴趣，'ICS'术语已经受到青睐，尽管这里将使用"CFC"。
CFC是细胞功能的测试，将淋巴细胞暴露于培养物中的抗原，然后测量任何诱导的细胞因子应答。处理测试培养物以用用荧光标记物标记的单克隆抗体染色细胞，并使用多聚甲醛化学固定细胞和净化样品。
CFC提供流式细胞术的功能，包括批量采样和多参数互相关，用于抗原特异性记忆反应的分析。使用CFC的研究者能够表型表征用测试抗原和表型亚型（例如，CD4 + 或CD8 + T细胞）培养的细胞，确定高于背景水平的产生频率的细胞因子。
与ELISPOT和Luminex方法相反，CFC可以关联来自特定的，表型 - 表征的细胞的多种细胞因子的产生。 ...
|
|
|
| T Follicular Helper Cell Coculture Assay
|
|
Author:
Date:
2014-01-05
[Abstract] T follicular helper (Tfh) cells constitute a distinct subset of CD4+ T cells specialized in providing help to B cells in germinal centers. Phenotypically, Tfh cells are characterized by their high expression of the chemokine receptor CXCR5 that allows their migration into B cell follicles as well as high expression of PD-1, BTLA, the co-stimulatory molecules ICOS and SLAM and the transcription factors BCL6 and cMaf. Tfh cells are the main producers of IL-21 as well as other cytokines like IL-4 and IL-10 critical for B cell survival and differentiation. Tfh cells drive somatic hypermutation and the generation of long-lived memory B cells and plasma cells having an essential role in the development of protective immunity. Developing a coculture system to measure the effects of ...
[摘要] T滤泡辅助（Tfh）细胞构成专门提供对生发中心中的B细胞的帮助的CD4 + T细胞的不同亚群。表型上，Tfh细胞的特征在于其高趋化因子受体CXCR5的表达，其允许它们迁移到B细胞滤泡以及PD-1，BTLA，共刺激分子ICOS和SLAM以及转录因子BCL6和cMaf的高表达。 Tfh细胞是IL-21以及其他细胞因子如IL-4和IL-10对B细胞存活和分化关键的主要生产者。 Tfh细胞驱动体细胞超突变和长寿记忆B细胞和浆细胞的产生在保护性免疫的发展中具有重要作用。开发共培养系统以测量Tfh细胞介导的B细胞帮助的效果是非常感兴趣的进一步我们的Tfh-B细胞相互作用的理解，并允许操纵文化条件调查潜在影响不同的微环境信号或配体/受体相互作用可能对Tfh细胞功能。
|
|
|