Silva Ana M, Oliveira Paulo J
MitoXT-Mitochondrial Toxicology and Experimental Therapeutics Laboratory, CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Cantanhede, Portugal.
Department of Life Sciences, University of Coimbra, Cantanhede, Portugal.
Methods Mol Biol. 2018;1782:7-29. doi: 10.1007/978-1-4939-7831-1_2.
In many studies, the evaluation of mitochondrial function is critical to understand how disease conditions or xenobiotics alter mitochondrial function. One of the classic end points that can be assessed is oxygen consumption, which can be performed under controlled yet artificial conditions. Oxygen is the terminal acceptor in the mitochondrial respiratory chain, namely, at an enzyme called cytochrome oxidase, which produces water in the process and pumps protons from the matrix to the intermembrane space. Several techniques are available to measure oxygen consumption, including polarography with oxygen electrodes or fluorescent/luminescent probes. The present chapter will deal with the determination of mitochondrial oxygen consumption by means of the Clark-type electrode, which has been widely used in the literature and still remains to be a reliable technique. We focus our technical description in the measurement of oxygen consumption by isolated mitochondrial fractions and by permeabilized cells.
在许多研究中，评估线粒体功能对于理解疾病状态或外源性物质如何改变线粒体功能至关重要。可以评估的经典终点之一是氧消耗，它可以在可控但人为的条件下进行。氧是线粒体呼吸链中的终端受体，即在一种名为细胞色素氧化酶的酶处，该过程中产生水并将质子从基质泵入膜间隙。有几种技术可用于测量氧消耗，包括使用氧电极的极谱法或荧光/发光探针。本章将介绍通过克拉克型电极测定线粒体氧消耗的方法，该方法在文献中已被广泛使用，并且仍然是一种可靠的技术。我们将技术描述重点放在分离的线粒体组分和透化细胞的氧消耗测量上。
