| Isolation of Nippostrongylus brasiliensis Larvae from Mouse Lungs
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Author:
Date:
2016-02-20
[Abstract] The rodent parasite Nippostrongylus brasiliensis (N. brasiliensis) models the salient features of helminth infection including skin penetration, migration from tissues to lung, maturation and egg production in the gut. As a potent activator of systemic and mucosal Th2 immune responses, Nippostrongylus brasiliensis has been extensively used to study host protective immunity and in vivo regulation of Th2 immune response. Six to eight week old C57Bl/6J, Balb/c mice or any other strains are suitable, as all are susceptible to infection. Inocula of 150-650 L3 larvae can be administered by subcutaneous injection, but for greatest consistency a dose of 550 L3 larvae is routinely used for experimental purposes. We have optimized three different protocols for ...
[摘要] 啮齿动物寄生虫（Nippostrongylus brasiliensis）（<巴西> ）模拟蠕虫感染的显着特征，包括皮肤渗透，从组织到肺的迁移，肠道中的成熟和产蛋。作为全身性和粘膜Th2免疫应答的有效激活剂，巴西尼罗巴戟天已广泛用于研究宿主保护性免疫和体内调节Th2免疫应答。 6至8周龄的C57Bl/6J，Balb/c小鼠或任何其它菌株是合适的，因为所有这些菌株都易于感染。 150-650LL幼虫的接种可以通过皮下注射给药，但是为了最大的一致性，通常将550LL幼虫的剂量用于实验目的。我们已经优化了三种不同的方案用于从感染了巴西尼罗巴戟甲L3阶段的小鼠的肺部分离幼虫。幼虫可以从体内任何部位接种后18-60小时迁移到肺部。在接种后48小时，出现在肺峰中的幼虫的数目，并且建议在48小时进行分离/收获以获得每种收获方法的最大一致性：
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| Isolation and Culture of Human Endometrial Epithelial Cells and Stromal Fibroblasts
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Author:
Date:
2015-10-20
[Abstract] Purification and culture of endometrial epithelial cells (eEC) and stromal fibroblasts (eSF) from endometrial biopsies allows for downstream cell-specific in vitro studies. The utility of this protocol is the ease with which cells are purified without contamination from unwanted cell types, and the ability to use patient-paired eEC and eSF in experiments. These methods have been previously published, but here the protocol has been updated for maximum efficiency.
[摘要] 来自子宫内膜活检的子宫内膜上皮细胞（eEC）和基质成纤维细胞（eSF）的纯化和培养允许下游细胞特异性体外研究。 该协议的效用是细胞被净化而不受不想要的细胞类型的污染，以及在实验中使用患者配对的eEC和eSF的能力。 这些方法以前已经发布，但在这里协议已经更新为最大的效率。
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| Determining the Relative Fitness Score of Mutant Viruses in a Population Using Illumina Paired-end Sequencing and Regression Analysis
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Author:
Date:
2015-05-20
[Abstract] Recent advances in DNA sequencing capacity to accurately quantify the copy number of individual variants in a large and diverse population allows in parallel determination of the phenotypic effects caused by each genetic modification. This systematic profiling approach is a combination of forward and reverse genetics, which we refer to as quantitative high-resolution genetics (qHRG). This protocol describes how to determine the relative fitness score of each variant compared to wild type (WT) virus based on its frequency determined by Illumina sequencing. Random mutagenesis techniques will be used to introduce randomization at each codon position of the targeted region, thereby generating a comprehensive input mutant library with substitutions at each position of interest (Qi et al. ...
[摘要] DNA测序能力的最近进展，准确量化大和多样群体中单个变体的拷贝数允许平行测定由每个遗传修饰引起的表型效应。这种系统分析方法是正向和反向遗传学的组合，我们称之为定量高分辨率遗传学（qHRG）。该方案描述了如何基于其通过Illumina测序确定的频率来确定每个变体与野生型（WT）病毒相比的相对适合度评分。随机诱变技术将用于在目标区域的每个密码子位置引入随机化，从而产生在每个感兴趣位置具有取代的综合输入突变文库（Qi等人，2014; Wu 等人，2014a; Wu ，2014b）。选择后，每个选定的文库将通过Illumina配对末端测序进行测序，并且确定每个突变的频率。基于频率的变化，可以用回归分析计算每个突变体的相对适合度分数。
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