| Biotinylation and Purification of Plasma Membrane-associated Proteins from Rodent Cultured Neurons
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Author:
Date:
2016-05-20
[Abstract] This protocol aims at the biotin labeling and affinity purification of plasma membrane proteins from cultured neurons. Protein biotinylation consists in the covalent attachment of biotin to proteins. Biotin is a membrane unpermeable molecule with a small size (MW 244.31 g/mol) and therefore does not interfere with the normal function of proteins. Biotin binds to streptavidin and avidin molecules with high affinity. This binding is extremely resistant to temperature, pH and proteolysis, which allows capture and purification of plasma membrane proteins. Moreover, proteins can bind several biotin molecules, that will allow the consequent binding of several streptavidin or avidin molecules, increasing the sensitivity of detection of the proteins of interest. In this protocol proteins at the ...
[摘要] 该方案旨在从培养的神经元生物素标记和亲和纯化质膜蛋白。 蛋白质生物素化包括生物素与蛋白质共价连接。 生物素是具有小尺寸（MW 244.31g / mol）的膜不可渗透分子，因此不干扰蛋白质的正常功能。 生物素以高亲和力结合链霉抗生物素蛋白和抗生物素蛋白分子。 这种结合对温度，pH和蛋白水解具有极高的抗性，可以捕获和纯化质膜蛋白。 此外，蛋白质可以结合几个生物素分子，这将允许几个链霉抗生物素蛋白或抗生物素蛋白分子的结合，增加感兴趣的蛋白质的检测灵敏度。 在该方案中，活的培养神经元的细胞表面的蛋白质被生物素化。 制备神经元提取物并用NeutrAvidin偶联的珠收集生物素化的蛋白质，并通过Western印迹进行分析。
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| ELISA Detection of Endogenous Serum Albumin in the Mouse Brain: A Measure of Extravasation Following Brain Injury
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Author:
Date:
2015-05-05
[Abstract] After stroke and brain contusion, serum proteins extravasate into nerve tissue through disrupted blood-brain barrier (BBB). Because extravasations of serum proteins result in vasogenic brain edema, serum albumin level in the brain is an indicator of BBB disruption and brain edema after brain insults. In this protocol, extravasation of endogenous albumin is measured in the damaged mouse brain, which would be valuable in the evaluation of vasogenic brain edema formation (Michinaga et al., 2014).
[摘要] 在中风和脑挫伤后，血清蛋白通过破裂的血脑屏障（BBB）外渗到神经组织中。 因为血清蛋白的外渗导致血管性脑水肿，脑中的血清白蛋白水平是脑损伤后BBB破坏和脑水肿的指标。 在该方案中，在损伤的小鼠脑中测量内源性白蛋白的外渗，这在评价血管性脑水肿形成中是有价值的（Michinaga等人，2014）。
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