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FinnpipetteTM F1 Multichannel Pipettes

Company: Thermo Fisher Scientific
Catalog#: 4661030N
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High-throughput Microscopic Analysis of Salmonella Invasion of Host Cells
Author:
Date:
2018-09-20
[Abstract]  Salmonella is a Gram-negative bacterium causing a gastro-enteric disease called salmonellosis. During the first phase of infection, Salmonella uses its flagella to swim near the surface of the epithelial cells and to target specific site of infection. In order to study the selection criteria that determine which host cells are targeted by the pathogen, and to analyze the relation between infecting Salmonella (i.e., cooperation or competition), we have established a high-throughput microscopic assay of HeLa cells sequentially infected with fluorescent bacteria. Using an automated pipeline of image analysis, we quantitatively characterized a multitude of parameters of infected and non-infected cells. Based on this, we established a predictive model that ... [摘要]  沙门氏菌是革兰氏阴性细菌,引起称为沙门氏菌病的胃肠疾病。在感染的第一阶段,沙门氏菌使用其鞭毛在上皮细胞表面附近游泳并靶向特定的感染部位。为了研究确定哪种宿主细胞被病原体靶向的选择标准,并分析感染沙门氏菌( ie ,合作或竞争)之间的关系,我们有建立了对荧光细菌依次感染的HeLa细胞的高通量显微镜检测。使用自动化图像分析管道,我们定量表征了感染和未感染细胞的众多参数。基于此,我们建立了一个预测模型,使我们能够识别宿主细胞易受感染的参数。我们发现宿主细胞易损性有两个来源:病原体诱导的细胞易感性从沙门氏菌摄取中出现并持续存在于感染过程的后期阶段;以及与细胞固有属性相关的宿主细胞固有的脆弱性,例如局部细胞拥挤和胆固醇含量。我们的方法基于形态学或分子宿主细胞参数预测单层上皮细胞中沙门氏菌感染的概率。在这里,我们提供了工作流程的详细描述,包括基于计算机的分析管道。我们的方法有可能应用于研究宿主 - 病原体相互作用的其他组合。

【背景】鼠伤寒沙门氏菌血清型鼠伤寒沙门氏菌通过摄入受污染的食物或水感染宿主,引起沙门氏菌病。一旦细菌到达肠道的远端回肠,它们就会侵入广泛的宿主细胞,包括肠上皮细胞(Watson和Holden,2010)。在宿主细胞入侵的第一阶段,沙门氏菌选择其目标,使用其鞭毛游泳并扫描上皮表面(Misselwitz et ...

GFP-Grb2 Translocation Assay Using High-content Imaging to Screen for Modulators of EGFR-signaling
Author:
Date:
2017-09-05
[Abstract]  High-content screening is a useful tool to understand complex cellular processes and to identify genes, proteins or small molecule compounds that modulate such pathways. High-content assays monitor the function of a protein or pathway by visualizing a change in an image-based readout, such as a change in the localization of a reporter protein. Examples of this can be the translocation of a fluorescently tagged protein from the cytoplasm to the nucleus or to the plasma membrane. One protein that is known to undergo such translocation is the Growth Factor Receptor-bound protein 2 (GRB2) that is recruited to the plasma membrane upon stimulation of a growth factor receptor and subsequently undergoes internalization. We have used GFP-tagged Grb2 previously to identify genes that are involved ... [摘要]  高含量筛选是了解复杂细胞过程和鉴定调节这种途径的基因,蛋白质或小分子化合物的有用工具。高含量测定法通过显现基于图像的读数的变化来监测蛋白质或途径的功能,例如报告蛋白的定位的变化。其实例可以是将荧光标记的蛋白质从细胞质转移到细胞核或质膜。已知发生这种易位的一种蛋白质是生长因子受体结合蛋白2(GRB2),其在刺激生长因子受体并且随后经历内化后被招募到质膜。我们以前用GFP标记的Grb2来鉴定涉及EGFR信号的基因(Petschnigg等,2017)。最终,该测定可以适应于cDNA表达克隆(Freeman等人,2012),并且可用于早期药物发现以鉴定调节或抑制EGFR信号传导和内化的化合物(Antczak和Djaballah,2016)。
【背景】生长因子受体的信号转导对细胞维持正常功能至关重要,因此需要严格控制。生长因子受体的信号转导通过外部配体(例如,表皮生长因子,EGF)与跨膜受体(例如表皮生长因子受体(EGFR))和下游信号级联的活化(Yao等人,2015 )。 EGFR-信号传导的关键调节因子是生长因子受体结合蛋白2(Grb2),其由两个SH3结构域的内部SH2(Src同源性2)结构域组成。 Grb2通过其SH2结构域结合磷酸化酪氨酸残基上的活化生长因子受体,从而将受体活化与SOS-Ras-MAPK(丝裂原活化蛋白激酶)信号级联偶联。 ...

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