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Date:
2014-01-20
[Abstract] Intracellular pH (pHi) is an important physiological determinant of enzyme activity and cellular function (Kurkdjian and Guern, 1989). All proteins depend on a tightly regulated pH to maintain their structure and function. Protonation–deprotonation events can dictate the charge of biological surfaces and are integral steps in many metabolic reactions (Casey et al., 2010). Moreover, the proton gradient across the mitochondrial membrane is used to generate cellular energy and support other mitochondrial processes. As a result, cells have developed multiple mechanisms to maintain a narrow range of pHi in response to extra- and intracellular fluctuations in pH (Orij et al., 2012). Here, we describe a protocol for pHi measurement in live cells ...
[摘要] 细胞内pH(pH )是酶活性和细胞功能的重要生理决定因素(Kurkdjian和Guern,1989)。所有蛋白质依赖于严格调节的pH以维持其结构和功能。质子化 - 去质子化事件可以决定生物表面的电荷,并且是许多代谢反应中不可或缺的步骤(Casey等人,2010)。此外,穿过线粒体膜的质子梯度用于产生细胞能量和支持其他线粒体过程。因此,细胞已经发展了多种机制以响应于pH的胞外和细胞内波动维持pH范围的窄范围(Orij等人,2012)。在这里,我们描述了在使用荧光显微镜和pH敏感染料2',7'-双 - (2-羧乙基)-5-(和-6-氨基己酸)的活细胞中pH pH测量的方案, ) - 羧基荧光素乙酰氧基甲基酯(BCECF-AM)。该方法最近用于测定细胞内pH变化对全局组蛋白乙酰化水平的影响(McBrian等人,2013)。
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