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DPBS, no calcium, no magnesium

DPBS,无钙,无镁

Company: Thermo Fisher Scientific
Catalog#: 14190144
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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 ...

Analysis of the Effect of Sphingomyelinase on Rubella Virus Infectivity in Two Cell Lines
Author:
Date:
2018-09-05
[Abstract]  Rubella is a mildly contagious disease characterized by low-grade fever and a morbilliform rash caused by the rubella virus (RuV). Viruses often use cellular phospholipids for infection. We studied the roles of cellular sphingomyelin in RuV infection. Treatment of cells with sphingomyelinase (SMase) inhibited RuV infection in rabbit kidney-derived RK13 cells and African green monkey (Cercopithecus aethiops) kidney-derived Vero cells. Our data further demonstrated that RuV used cellular sphingomyelin and cholesterol for its binding to cells and membrane fusion at the step of virus entry. Detailed protocols of our assays, which assess the effects of SMase treatment on RuV infectivity in RK13 and Vero cells, are described. [摘要]  风疹是一种轻度传染性疾病,其特征是低风热和由风疹病毒(RuV)引起的麻疹样皮疹。 病毒通常使用细胞磷脂进行感染。 我们研究了细胞鞘磷脂在RuV感染中的作用。 用鞘磷脂酶(SMase)处理细胞抑制兔肾衍生的RK13细胞和非洲绿猴( Cercopithecus aethiops )肾源性Vero细胞中的RuV感染。 我们的数据进一步证明RuV在病毒进入的步骤中使用细胞鞘磷脂和胆固醇来结合细胞和膜融合。 描述了我们的测定的详细方案,其评估了SMase处理对RK13和Vero细胞中RuV感染性的影响。

【背景】风疹病毒(RuV)是一种正链RNA病毒,属于 Togaviridae 家族中的 Rubivirus 属。该家族有两个属, Rubivirus 和 Alphavirus 。风疹病毒是属的鞋底构件的风疹病毒,而许多病毒,例如塞姆利基森林病毒(SFV)和辛德毕斯病毒(SINV),是归类于甲病毒属。 RuV是风疹和先天性风疹综合征(CRS)的致病因子。风疹的特征是低烧,麻疹样皮疹和淋巴结肿大。它通常是一种轻微的疾病。然而,CRS是一种严重的疾病。 CRS导致在怀孕早期患有风疹的母亲所生的新生儿出现多器官缺陷。白内障,感音神经性听力损失和心血管缺陷在CRS中很常见。

以前的研究表明,细胞膜脂质作为RuV感染的结合或进入因子(Mastromarino ...

CRISPR/Cas Gene Editing of a Large DNA Virus: African Swine Fever Virus
Author:
Date:
2018-08-20
[Abstract]  Gene editing of large DNA viruses, such as African swine fever virus (ASFV), has traditionally relied on homologous recombination of a donor plasmid consisting of a reporter cassette with surrounding homologous viral DNA. However, this homologous recombination resulting in the desired modified virus is a rare event. We recently reported the use of CRISPR/Cas9 to edit ASFV. The use of CRISPR/Cas9 to modify the African swine fever virus genome resulted in a fast and relatively easy way to introduce genetic changes. To accomplish this goal we first infect primary swine macrophages with a field isolate, ASFV-G, and transfect with the CRISPR/Cas9 donor plasmid along with a plasmid that will express a specific gRNA that targets our gene to be deleted. By inserting a reporter cassette, we are ... [摘要]  大型DNA病毒(例如非洲猪瘟病毒(ASFV))的基因编辑传统上依赖于由报道盒组成的供体质粒与周围同源病毒DNA的同源重组。然而,这种导致所需修饰病毒的同源重组是罕见的事件。我们最近报道了使用CRISPR / Cas9编辑ASFV。使用CRISPR / Cas9修饰非洲猪瘟病毒基因组导致了引入遗传变化的快速且相对简单的方法。为了实现这一目标,我们首先用田间分离株ASFV-G感染原代猪巨噬细胞,并用CRISPR / Cas9供体质粒转染质粒,该质粒将表达靶向我们基因的特异性gRNA被删除。通过插入报告盒,我们能够通过有限稀释和噬菌斑纯化从亲本中纯化我们的重组病毒。我们以前曾报道将传统的同源重组方法与CRISPR / Cas9进行比较,结果导致重组增加超过4个对数。

【背景】 非洲猪瘟(ASF)是一种由ASF病毒(ASFV)引起的高度致命的猪传染性病毒性疾病。 ASFV的基因组由大约180-190千碱基对的双链DNA基因组组成。 ASFV引起一系列疾病,从高度致命到亚临床,取决于宿主特征和病毒株(Tulman et al。,2009)。 ASFV没有商业疫苗;实验上,2007年格鲁吉亚爆发的唯一能够抵御目前流行病毒株的疫苗(ASFV-G)是含有一个或多个病毒基因组缺失的减毒活疫苗,例如:( O'Donnell et ...

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