| Separation of Intracellular Vesicles for Immunoassays
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Author:
Date:
2015-08-20
[Abstract] The endosome/lysosome systems play important roles in immune cell functions as signaling platforms. Immune cells utilize these endosome/lysosome for signal transduction or intercellular communication to elicit the proper immune responses, regulating the localization or the association of the signaling complexes. Here we introduce the procedures to separate the intracellular vesicles such as endosomes or lysosomes, which could be useful to identify the subcellular localization of the signaling complexes.
[摘要] 内体/溶酶体系统在作为信号平台的免疫细胞功能中起重要作用。 免疫细胞利用这些内体/溶酶体进行信号转导或细胞间通讯，以引发适当的免疫应答，调节信号复合物的定位或缔合。 在这里我们介绍分离细胞内囊泡如内体或溶酶体的程序，这可能有助于识别信号复合物的亚细胞定位。
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| Preparation of Parasite Protein Extracts and Western Blot Analysis
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Author:
Date:
2014-06-05
[Abstract] In order to prepare protein extracts of Plasmodium falciparum blood stages for western blot analysis, infected red blood cells (iRBC) need to be separated from uninfected red blood cells (uRBC) which make up the bulk of the parasite culture. Depending on the localisation of the parasite protein of interest, different methods are available to achieve this. If the protein is present within the parasite or is attached to a cellular structure of the iRBC cell, saponin can be used. This reagent lyses the membranes of infected and uninfected erythrocytes, the Maurer´s clefts (vesicular structures in the iRBC) and the parasitophorous vacuole membrane containing the parasite but leaves the parasite plasma membrane intact, providing a convenient procedure to isolate intact parasites ...
[摘要] 为了制备用于蛋白质印迹分析的恶性疟原虫血液阶段的蛋白质提取物，需要将感染的红细胞（iRBC）与构成寄生虫的主体的未感染的红细胞（uRBC）分离文化。根据感兴趣的寄生虫蛋白的定位，可以使用不同的方法来实现这一点。如果蛋白质存在于寄生虫内或附着于iRBC细胞的细胞结构，则可以使用皂苷。该试剂裂解感染的和未感染的红细胞的膜，Maurer's裂口（iRBC中的囊泡结构）和含有寄生虫的寄生虫液泡膜，但是使寄生虫质膜完整，提供了一种方便的方法来分离没有uRBC的完整寄生虫。然而，该方法的缺点是iRBC的宿主细胞胞质和寄生泡（PV）含量丢失。如果这必须避免，可以使用Percoll梯度将完整的iRBC与uRBC分离。然后可以使用Tetanolysin和皂苷的顺序处理来从寄生虫选择性释放iRBC细胞溶质和PV含量。这些选择性裂解方法也适合于确定目标蛋白质的亚细胞定位
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