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Absorbance Reader

ELx808吸光度读数器

Company: BioTek Instruments
Catalog#: ELx808
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Preparation of Mosquito Salivary Gland Extract and Intradermal Inoculation of Mice
Author:
Date:
2017-07-20
[Abstract]  Mosquito-transmitted pathogens are among the leading causes of severe disease and death in humans. Components within the saliva of mosquito vectors facilitate blood feeding, modulate host responses, and allow efficient transmission of pathogens, such as Dengue, Zika, yellow fever, West Nile, Japanese encephalitis, and chikungunya viruses, as well as Plasmodium parasites, among others. Here, we describe standardized methods to assess the impact of mosquito-derived factors on immune responses and pathogenesis in mouse models of infection. This protocol includes the generation of mosquito salivary gland extracts and intradermal inoculation of mouse ears. Ultimately, the information obtained from using these techniques can help reveal fundamental mechanisms of interaction between ... [摘要]  蚊子传播的病原体是人类严重疾病和死亡的主要原因之一。蚊子唾液中的组分促进血液供体,调节宿主反应,并允许有效传播病原体,如登革热,紫草,黄热病,西尼罗河,日本脑炎和基孔肯雅病毒,以及疟原虫寄生虫等。在这里,我们描述了评估蚊子衍生因子对小鼠感染模型中免疫反应和发病机制的影响的标准化方法。该方案包括产生蚊子唾液腺提取物和皮内接种小鼠耳朵。最终,使用这些技术获得的信息可以帮助揭示病原体,蚊子和哺乳动物宿主之间的相互作用的基本机制。此外,该协议还可以帮助建立疫苗或治疗剂的临床前检测改进的感染挑战模型,以考虑通过蚊子传播的自然途径。
【背景】在探测血液的同时,蚊子接种有助于喂养的唾液,但如果蚊子曾经感染过感染个体,也可能含有病原体。蚊子唾液在建立感染,促进传播,调节免疫应答和加剧西尼罗河病毒发病过程中发挥重要作用(Schneider等人,2006; Styer等人)。 ,2011),登革热病毒(Cox等人,2012; Conway等人,2014; McCracken等人,2014; Schmid ,2016),基孔肯雅病毒(Agarwal等人,2016),Semliki Forest病毒(Pingen等人,2016) ,裂谷热病毒(Le Coupanec等人,2013)和疟原虫寄生虫(Schneider等人,2011)感染。许多重要问题仍然存在,需要改进动物模型。
 而通过感染的蚊子接种最好地模仿自然传播,接种剂量的高度变异性和有限的昆虫设施的可用性导致这种程序的有限使用。此外,使用感染的蚊子时,不能控制唾液的含量和蚊子组分的存在或不存在。作为替代方案,未感染的雌性蚊子的“斑点喂养”,然后通过针对病原体进行真皮内接种,模拟了唾液在小鼠皮肤中的自然沉积并递送了一定剂量的病原体。 ...

Assay to Evaluate Vascular Permeability Induction in Mice
Author:
Date:
2013-11-20
[Abstract]  Dengue virus infection usually courses as a benign self-limited fever, called dengue fever. However, on occasions it can progress to a life-threatening complication known as severe dengue (SD). A hallmark of SD is a sharp increase in vascular permeability. Secondary infections are considered a risk factor to develop SD, presumably through a mechanism called Antibody-Dependent Enhancement (ADE) of infection in cells with the capacity to bind antigen-antibody complexes, such as macrophages, and to trigger a subsequent aberrant cytokine response. The massive release of cytokine from macrophages has been postulated to cause changes in vascular permeability. The vascular permeability assay presented in this protocol is designed to assess whether any compound or cell-secreted product ... [摘要]  登革热病毒感染通常称为良性自限发热,称为登革热。然而,偶尔,它可以进展为严重登革热(SD)的危及生命的并发症。 SD的标志是血管通透性的急剧增加。二次感染被认为是发展SD的风险因素,可能是通过在具有结合抗原 - 抗体复合物(例如巨噬细胞)能力的细胞中感染的感染的机制,称为抗体依赖性增强(ADE)引发随后的异常细胞因子应答。已经假定巨噬细胞中细胞因子的大量释放导致血管通透性的变化。在该方案中呈现的血管渗透性测定设计为评估来自患者的血清中存在的任何化合物或细胞分泌的产物或可溶性因子是否可以在小鼠中诱导血浆渗漏。在实验室中使用该测试以确定在存在促进性抗体的情况下由登革热病毒或登革热病毒感染的巨噬细胞在体外产生的细胞因子和可溶性因子是否能够在体内诱导血浆渗漏。在存在促进抗体的情况下,将巨噬细胞用登革病毒或登革热病毒感染48小时。此后,收集含有由巨噬细胞释放的细胞因子和可溶性因子的条件上清液,并腹膜内接种到CD-1小鼠中。在第一次接种后二十四小时,用伊文思蓝染料以第二剂量再次接种小鼠。再过24小时后,对小鼠实施安乐死,通过分光光度分析测定血液和肺中存在的伊文思蓝的量。该测定能够显示条件培养基在接种的动物中诱导血管通透性改变的能力的差异(Puerta-Guardo等人,2013)。

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