{{'Search' | translate}}
 

Fetal Bovine Serum

胎牛血清

Company: Thermo Fisher Scientific
Catalog#: 10270106
Bio-protocol()
Company-protocol()
Other protocol()

Isolation and Culture of Mouse Lung ILC2s
Author:
Date:
2018-10-05
[Abstract]  Group 2 Innate Lymphoid Cells (ILC2) play an important role in immune responses at barrier surfaces, notably in the lung during airway allergic inflammation or asthma. Several studies have described methods to isolate ILC2s from wild-type naive mice, most of them using cell sorting to obtain a pure population. Here, we describe in detail, a simple, efficient method for isolation and culture of lung mouse ILC2s. Lungs from Rag2-/- mice pretreated with IL-33 are collected and processed into single cell suspensions. Lymphoid cells are then recovered by density gradient separation. Lin-CD45+ cells are selected by depletion of lineage positive cells followed by positive selection of CD45+ cells. Culture of the isolated cells for several days ... [摘要]  第2组先天性淋巴细胞(ILC2)在屏障表面,特别是在气道过敏性炎症或哮喘期间的肺中的免疫应答中起重要作用。一些研究已经描述了从野生型幼稚小鼠中分离ILC2的方法,其中大多数使用细胞分选来获得纯种群。在这里,我们详细描述了一种简单有效的肺小鼠ILC2分离和培养方法。收集用IL-33预处理的 Rag2 - / - 小鼠的肺并加工成单细胞悬浮液。然后通过密度梯度分离回收淋巴样细胞。通过耗尽谱系阳性细胞然后阳性选择CD45 + 细胞来选择Lin - CD45 + 细胞。将分离的细胞培养数天导致高度纯化的ILC2群体表达典型的细胞表面标志物(CD90.2,Sca1,CD25,CD127和IL-33R)。这些细胞可在培养物中扩增长达10天,并用于多种离体测定或体内过继转移实验。
【背景】第2组先天性淋巴细胞(ILC2)是组织驻留细胞,其在抗寄生虫先天免疫以及过敏性炎症的发展中起关键作用。它们通过产生大量的2型细胞因子IL-5和IL-13对上皮细胞衍生的细胞因子如白细胞介素-33(IL-33)起反应,后者又诱导嗜酸性粒细胞增多和粘液产生(Cayrol和Girard,2018)。为了更好地表征这些细胞的功能和调节,许多组通过荧光激活细胞分选(FACS)从野生型小鼠(WT)的肺中分选ILC2。由于稳定状态下肺中存在的ILC2数量较少,因此该方法导致纯化细胞的产量较低(每只小鼠1×10 ...

Zebrafish Embryo Xenograft and Metastasis Assay
Author:
Date:
2018-09-20
[Abstract]  Xenograft models, and in particular the mouse xenograft model, where human cancer cells are transplanted into immunocompromised mice, have been used extensively in cancer studies. Although these models have contributed enormously to our understanding of cancer biology, the zebrafish xenograft model offers several advantages over the mouse model. Zebrafish embryos can be easily cultured in large quantities, are small and easy to handle, making it possible to use a high number of embryos for each experimental condition. Young embryos lack an efficient immune system. Therefore the injected cancer cells are not rejected, and the formation of primary tumors and micrometastases is rapid. Transparency of the embryos enables imaging of primary tumors and metastases in an intact and living embryo. ... [摘要]  异种移植模型,特别是小鼠异种移植模型,其中人癌细胞被移植到免疫受损小鼠中,已广泛用于癌症研究。尽管这些模型对我们对癌症生物学的理解做出了巨大贡献,但斑马鱼异种移植模型与鼠标模型相比具有多种优势。斑马鱼胚胎可以很容易地大量培养,体积小且易于处理,因此可以在每种实验条件下使用大量胚胎。年轻的胚胎缺乏有效的免疫系统。因此,注射的癌细胞不被排斥,并且原发性肿瘤和微转移的形成是快速的。胚胎的透明度使得能够在完整和活的胚胎中对原发性肿瘤和转移瘤进行成像。在这里,我们描述了一种方法,其中表达GFP的肿瘤细胞被注射到斑马鱼胚胎的心包空间。在注射后4天,对胚胎成像并分析原发性肿瘤和远端微转移的形成。

【背景】 斑马鱼( Danio rerio )是一种小型淡水鱼,不仅在发育生物学中,而且在生物医学中越来越受欢迎。快速发展的领域之一是使用斑马鱼作为癌症生物学研究的模型。斑马鱼高效可靠地交配,可以产生大量小,透明,外部发育的后代,可以在例如,多孔板中轻松培养(White et al。 >,2013)。有趣的是,人类肿瘤细胞可以植入斑马鱼胚胎中,许多异种移植物能够通过在胚胎中生长,侵入和转移来忠实地重演它们的恶性行为(Lee et al。,2005; Nicoli et al。,2007; Chapman et ...

A Quantitative Heterokaryon Assay to Measure the Nucleocytoplasmic Shuttling of Proteins
Author:
Date:
2018-09-05
[Abstract]  Many proteins appear exclusively nuclear at steady-state but in fact shuttle continuously back and forth between the nucleus and the cytoplasm. For example, nuclear RNA-binding proteins (RBPs) often accompany mRNAs to the cytoplasm, where they can regulate subcellular localization, translation and/or decay of their cargos before shuttling back to the nucleus. Nucleocytoplasmic shuttling must be tightly regulated, as mislocalization of several RBPs with prion-like domains such as FUS and TDP-43 causes the cytoplasmic accumulation of solid pathological aggregates that have been implicated in neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Traditionally, interspecies heterokaryon assays have been used to determine whether a nuclear ... [摘要]  许多蛋白质在稳态下仅出现核,但事实上在细胞核和细胞质之间连续地来回穿梭。例如,核RNA结合蛋白(RBP)通常伴随mRNA到达细胞质,在那里它们可以在穿梭回到细胞核之前调节其货物的亚细胞定位,翻译和/或腐烂。必须严格调节核质穿梭,因为几种RBP与朊病毒样结构域如FUS和TDP-43的错误定位导致固体病理性聚集体的细胞质积累,这些聚集体与肌萎缩侧索硬化症(ALS)和额颞叶痴呆等神经退行性疾病有关。 (FTD)。传统上,种间异核体分析已被用于确定感兴趣的核蛋白是否穿梭;这些分析是基于来自两个不同物种(例如,小鼠和人类)的供体和受体细胞之间的融合,可以根据不同的染色质染色模式区分,并检测蛋白质的外观。受体核。然而,异核体的鉴定需要经验并且容易出错,这使得难以获得用于定量研究的高质量数据。此外,荧光标记的RBP在供体细胞中的瞬时过表达通常导致其异常的亚细胞定位。在这里,我们提出定量测定,其中表达接近生理水平的eGFP标记的RBP的稳定供体细胞系与表达膜标记物CAAX-mCherry的受体细胞融合,允许容易地鉴定和成像大量高可信度异核体。我们的测定法可用于测量任何感兴趣的核蛋白在不同细胞类型,不同细胞条件下或突变蛋白之间的穿梭活性。

【背景】要了解蛋白质的各种功能,重要的是找出它在细胞内定位的位置。标准的微观和生物化学方法仅在其稳态浓度高于检测阈值时才揭示蛋白质的存在。他们不排除它在短暂地定位的情况下扮演其他重要角色的可能性(Gama-Carvalho和Carmo-Fonseca,2001)。例如,许多RBP在不同的细胞区室中发挥作用,它们伴随着它们的结合mRNA(通常未检测到)并连接真核基因表达的多个步骤(Müller-McNicoll和Neugebauer,2013)。 ...

Comments