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Manual micromanipulator

Company: World Precision Instruments
Catalog#: M3301
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Brain-localized and Intravenous Microinjections in the Larval Zebrafish to Assess Innate Immune Response
Author:
Date:
2021-04-05
[Abstract]  

Creating a robust and controlled infection model is imperative for studying the innate immune response. Leveraging the particular strengths of the zebrafish model system, such as optical transparency, ex utero development, and large clutch size, allows for the development of methods that yield consistent and reproducible results. We created a robust model for activation of innate immunity by microinjecting bacterial particles or live bacteria into larval zebrafish, unlike previous studies which largely restricted such manipulations to embryonic stages of zebrafish. The ability to introduce stimuli locally or systemically at larval stages provides significant advantages to examine host response in more mature tissues as well as the possibility to interrogate adaptive immunity at older

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[摘要]  [摘要]创建一个健壮和控制感染模型是势在必行用于学习的先天免疫应答。凭借特殊的斑马鱼模型系统的优势,比如光学透明性,前子宫发展,以及大离合器的尺寸,允许小号的了的,其产生方法的发展是一致和可重复的 结果。我们通过将细菌颗粒或活细菌显微注射到幼虫斑马鱼中创建了一个强大的激活先天免疫的模型,这与以前的研究在很大程度上将此类操作限制在斑马鱼的胚胎阶段不同。在幼虫阶段局部或全身引入刺激的能力提供了显着的优势,可检查更成熟组织中的宿主反应,并在较老的幼虫阶段询问适应性免疫的可能性。这个协议描述2种显微注射的不同模式以引入脂多糖(LPS)或细菌到活幼虫斑马鱼:一个本地化到的脑,和另一个为对经由尾静脉血流丛。


图形摘要:

示意性示出了两种不同的模式的幼虫斑马鱼显微注射,无论是在脑实质或在血流中静脉内。ř eagents引入斑马鱼,以评估免疫应答中作为在协议中描述的“注入组件”被描绘。

[背景]感染过程中复杂的相互作用需要使用体内动物模型来充分了解病原体与其宿主之间的动态相互作用。研究此现象需要病原体传递的受控和可靠方法。斑马鱼已被用作研究多种病原体免疫应答的模型(Menudier等,1996; Davis等,2002; Neely等,2002; Prouty等,2003; van der ...

Liposomal Clodronate-mediated Macrophage Depletion in the Zebrafish Model
Author:
Date:
2021-03-20
[Abstract]  

The ability to conduct in vivo macrophage-specific depletion remains an effective means to uncover functions of macrophages in a wide range of physiological contexts. Compared to the murine model, zebrafish offer superior imaging capabilities due to their optical transparency starting from a single-cell stage to throughout larval development. These qualities become important for in vivo cell specific depletions so that the elimination of the targeted cells can be tracked and validated in real time through microscopy. Multiple methods to deplete macrophages in zebrafish are available, including genetic (such as an irf8 knockout), chemogenetic (such as the nitroreductase/metronidazole system), and toxin-based depletions (such as using clodronate liposomes). The use of clodronate-containing

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[摘要]  [摘要]为了进行能力在体内巨噬细胞比耗尽仍然是有效的手段,在广泛的生理环境的巨噬细胞的揭开功能。与鼠模型相比,斑马鱼具有良好的成像能力,因为它们的光学透明性从单细胞阶段到整个幼体发育期开始。这些素质成为重要的体内细胞特异的枯竭,使靶细胞的消除可以跟踪并通过显微镜进行实时验证。多种方法以耗尽巨噬细胞在斑马鱼中是可用的,包括遗传(如IRF8敲除),chemogenetic(如在 硝基还原酶/甲硝唑系统)和基于毒素的消耗(例如使用氯膦酸盐脂质体)。在吞噬脂质体后,使用含氯膦酸盐的脂质体诱导巨噬细胞凋亡可有效地消耗巨噬细胞并测试其吞噬能力。在这里,我们描述了通过静脉注射补充有荧光右旋糖酐共轭物的脂质体氯膦酸盐对斑马鱼幼虫体内巨噬细胞进行系统耗竭的详细方案。与荧光右旋糖酐共注射可以实时跟踪巨噬细胞耗竭情况,从验证静脉内注射成功到摄取巨噬细胞分子及其最终死亡开始。为了验证高度的巨噬细胞耗竭,可以在幼虫早期进行氯膦酸盐注射时通过快速中性红色活体染料染色来确定脑巨噬细胞(小胶质细胞)的清除水平。

图形概要:

幼虫斑马鱼脂质体氯膦酸盐体内巨噬细胞特异性清除的实验工作流程


[背景]巨噬细胞是先天免疫系统的关键组成部分,并响应于感染,无菌性炎症,与环境变化中发挥重要作用。使巨噬细胞功能与不同生理环境中相互作用细胞类型的复杂混合物脱钩的最有效方法之一是能够特异性消除巨噬细胞并分析表型结果。小鼠中的此类耗竭实验为巨噬细胞的作用提供了很多见识(Hua等人,2018; ...

Expression and Analysis of Flow-regulated Ion Channels in Xenopus Oocytes
Author:
Date:
2017-04-20
[Abstract]  Mechanically-gated ion channels play key roles in mechanotransduction, a process that translates physical forces into biological signals. Epithelial and endothelial cells are exposed to laminar shear stress (LSS), a tangential force exerted by flowing fluids against the wall of vessels and epithelia. The protocol outlined herein has been used to examine the response of ion channels expressed in Xenopus oocytes to LSS (Hoger et al., 2002; Carattino et al., 2004; Shi et al., 2006). The Xenopus oocyte is a reliable system that allows for the expression and chemical modification of ion channels and regulatory proteins (George et al., 1989; Palmer et al., 1990; Sheng et al., 2001; Carattino et al., 2003). ... [摘要]  机械门控离子通道在机械传导中起关键作用,这是将物理力量转化为生物信号的过程。 上皮细胞和内皮细胞暴露于层流剪切应力(LSS),这是通过流体流向血管壁和上皮细胞壁所施加的切向力。 本文概述的方案已用于检查在非洲爪蟾卵母细胞中表达的离子通道对LSS的反应(Hoger等,2002; Carattino等,2004; Shi等,2006)。 非洲爪蟾卵母细胞是允许离子通道和调节蛋白的表达和化学修饰的可靠系统(George等,1989; Palmer等,1990; Sheng等,2001; Carattino等,2003)。 因此,该技术适用于研究允许流激活通道响应LSS的分子机制。
【背景】排列血管的泌尿道和内皮细胞的上皮细胞经受移动流体引起的机械力。这些力是层流剪切应力(LSS),与管状结构壁相切的摩擦力,以及垂直于流动方向的周向拉伸。令人信服的证据表明,LSS是响应肾和血管管状结构的流动变化而观察到的生理反应的主要决定因素(Satlin等,2001; Liu等,2003; Weinbaum等,2010) 。在这些设置中,离子通道具有将流体剪切应力传递到生物信号中的重要作用(Ranade等,2015)。例如,在肾的远端肾单位中,Na +再吸收和K ...

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