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MEM Non-Essential Amino Acids Solution (100X)

MEM非必需氨基酸溶液,100X

Company: Thermo Fisher Scientific
Catalog#: 11140050
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HCV Reporter System (Viral Infection-Activated Split-Intein-Mediated Reporter System) for Testing Virus Cell-to-cell Transmission ex-vivo
Author:
Date:
2018-08-05
[Abstract]  Hepatitis C virus (HCV) spread involves two distinct entry pathways: cell-free transmission and cell-to-cell transmission. Cell-to-cell transmission is not only an efficient way for viruses to spread but also an effective method for escaping neutralizing antibodies. We adapted the viral infection-activated split-intein-mediated reporter system (VISI) and developed a straightforward model for Live-cell monitoring of HCV cell-to-cell transmission ex-vivo: co-culture of HCV infected donor cells (red signal) with uninfected recipient cells (green signal) and elimination of the cell-free transmission by adding potent neutralizing antibody AR3A in the supernatant. With this model, the efficiency of cell-to-cell transmission can be evaluated by counting the number of foci designated by ... [摘要]  丙型肝炎病毒(HCV)传播涉及两种不同的进入途径:无细胞传播和细胞间传播。 细胞间传播不仅是病毒传播的有效方式,也是逃避中和抗体的有效方法。 我们采用了病毒感染激活的分裂 - 内含肽介导的报告系统(VISI),并开发了一种直接模型,用于活细胞监测HCV细胞间传递离体:共培养 HCV感染的供体细胞(红色信号)与未感染的受体细胞(绿色信号)和通过在上清液中加入有效的中和抗体AR3A消除无细胞的传递。 利用该模型,可以通过计数受体细胞的绿色信号指定的病灶数来评估细胞间传递的效率。

【背景】越来越多的证据证明病毒可以在受感染的组织中使用不同的传播途径(Sattentau,2008; Zhong et al。,2013)。对于HCV传播,无细胞传播和细胞间传播均可介导肝细胞之间的病毒转移。虽然无细胞传播引发HCV感染,但认为细胞 - 细胞传递直接将HCV转移至相邻的肝细胞。它提供了抵抗中和抗体并有助于病毒持久性的极好方法(Brimacombe et al。,2011; Xiao et al。,2014)。之前的文章也证明了一些促进细胞传递的宿主因子,如清道夫受体BI(SR-BI),CD81,紧密连接蛋白claudin-1(CLDN1),Occludin(OCLN),表皮生长因子受体(EGFR)。 (Witteveldt et al。,2009; ...

Preserve Cultured Cell Cytonemes through a Modified Electron Microscopy Fixation
Author:
Date:
2018-07-05
[Abstract]  Immunocytochemistry of cultured cells is a common and effective technique for determining compositions and localizations of proteins within cellular structures. However, traditional cultured cell fixation and staining protocols are not effective in preserving cultured cell cytonemes, long specialized filopodia that are dedicated to morphogen transport. As a result, limited mechanistic interrogation has been performed to assess their regulation. We developed a fixation protocol for cultured cells that preserves cytonemes, which allows for immunofluorescent analysis of endogenous and over-expressed proteins localizing to the delicate cellular structures. [摘要]  培养细胞的免疫细胞化学是用于确定细胞结构内蛋白质的组成和定位的常用且有效的技术。 然而,传统的培养细胞固定和染色方案不能有效地保存培养的细胞色素,长期专门用于形态发生转运的丝状伪足。 结果,进行了有限的机械审讯以评估其监管。 我们开发了一种用于培养细胞的固定方案,该方案保留了细胞质,允许对内源性和过表达的蛋白质进行免疫荧光分析,这些蛋白质定位于脆弱的细胞结构。

【背景】Cytonemes被分类为薄的(~200nm直径)基于肌动蛋白的丝状伪足,长度超过2μm,可以转运形态发生素(Ramírez-Weber和Kornberg,1999)。这些信号结构首先在发育中的 Drosophila 翼成像盘中进行了详细分类和描述,随后在小鼠,小鸡和斑马鱼模型生物中进行了观察(Ramírez-Weber和Kornberg,1999; Sanders et al。,2013; Stanganello et al。,2015)。在大多数情况下,只有对过表达的荧光标记蛋白进行实时成像才能进行细胞色素检测。由于传统的固定方案未能保存这些脆弱的细丝,因此对培养细胞的细胞色素的检查受到限制。这些并发症一直是决定在发育和组织稳态期间驱动细胞色素形成和功能的细胞机制以及确定这些过程是否在疾病中被破坏的限制因素。

为了克服这些限制,我们开发了一种基于修饰电子显微镜固定剂(MEM-fix)的方案,该方案可以保留培养细胞的细胞质。 ...

Enhancement of Mucus Production in Eukaryotic Cells and Quantification of Adherent Mucus by ELISA
Author:
Date:
2018-06-20
[Abstract]  The mucosal surfaces of the gastrointestinal, respiratory, reproductive, and urinary tracts, and the surface of the eye harbor a resident microflora that lives in symbiosis with their host and forms a complex ecosystem. The protection of the vulnerable epithelium is primarily achieved by mucins that form a gel-like structure adherent to the apical cell surface. This mucus layer constitutes a physical and chemical barrier between the microbial flora and the underlying epithelium. Mucus is critical to the maintenance of a homeostatic relationship between the microbiota and its host. Subtle deviations from this dynamic interaction may result in major implications for health. The protocol in this article describes the procedures to grow low mucus-producing HT29 and high mucus-producing ... [摘要]  胃肠道,呼吸道,生殖道和泌尿道的粘膜表面以及眼睛表面都有一个居住的微生物群落,它们与宿主共生并形成一个复杂的生态系统。脆弱的上皮细胞的保护主要通过形成附着于顶端细胞表面的凝胶样结构的粘蛋白实现。该粘液层构成了微生物菌群和下层上皮之间的物理和化学屏障。粘液对维持微生物群与宿主之间的稳态关系至关重要。与这种动态互动的细微差异可能会对健康产生重大影响。本文中的方案描述了生长低粘液产生HT29和高粘液产生HT29-MTX-E12细胞的程序,维持细胞并通过ELISA将其用于粘液定量。此外,还介绍了如何评估分泌黏液的数量。该系统可用于研究粘液对抗细菌毒素的保护作用,例如测试不同培养条件对粘液产生的影响或分析分子通过粘液层的扩散。由于本方案中使用的ELISA可用于不同的物种和粘液蛋白,因此也可以使用其他细胞类型。

【背景】身体与外部环境的界面由粘膜表面形成。这些粘膜上皮组织可以在例如胃肠道,呼吸道,生殖道和尿道以及眼睛表面发现。由于它们暴露于外部环境中,许多微生物会聚集在这些组织中。因此,这些上皮细胞已经进化出多种防御机制来回应其易受微生物攻击的影响。许多防御性化合物被分泌到粘膜液中,包括粘蛋白,抗体,防御素,protegrin,聚集蛋白,cathelicidins,溶菌酶,组蛋白和一氧化氮(Kagnoff和Eckmann,1997,Lu等人,2002 ...

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