Mitochondria serve several important roles in maintaining cellular homeostasis, including adenosine triphosphate (ATP) synthesis, apoptotic signalling, and regulation of both reactive oxygen species (ROS) and calcium. Therefore, mitochondrial studies may reveal insights into metabolism at higher levels of physiological organization. The apparent complexity of mitochondrial function may be daunting to researchers new to mitochondrial physiology. This review is aimed, therefore, at such researchers to provide a brief, yet approachable overview of common techniques used to assess mitochondrial function. Here we discuss the use of high-resolution respirometry in mitochondrial experiments and common analytical platforms used for this technique. Next, we compare the use of common mitochondrial preparation techniques, including adherent cells, tissue homogenate, permeabilized fibers and isolated mitochondria. Finally, we outline additional techniques that can be used in tandem with high-resolution respirometry to assess additional aspects of mitochondrial metabolism, including ATP synthesis, calcium uptake, membrane potential and reactive oxygen species emission. We also include limitations to each of these techniques and outline recommendations for experimental design and interpretation. With a general understanding of methodologies commonly used to study mitochondrial physiology, experimenters may begin contributing to our understanding of this organelle, and how it affects other physiological phenotypes.

线粒体在维持细胞稳态中发挥着多种重要作用，包括三磷酸腺苷 (ATP) 合成、细胞凋亡信号传导以及活性氧 (ROS) 和钙的调节。因此，线粒体研究可能揭示更高水平的生理组织的新陈代谢。线粒体功能的明显复杂性可能会让刚接触线粒体生理学的研究人员感到畏惧。因此，本综述旨在为这些研究人员提供用于评估线粒体功能的常用技术的简短但易于理解的概述。在这里，我们讨论高分辨率呼​​吸测定法在线粒体实验中的使用以及用于该技术的常见分析平台。接下来，我们比较常见线粒体制备技术的使用，包括贴壁细胞、组织匀浆、透化纤维和分离线粒体。最后，我们概述了可与高分辨率呼​​吸测量法结合使用的其他技术，以评估线粒体代谢的其他方面，包括 ATP 合成、钙吸收、膜电位和活性氧排放。我们还包括每种技术的局限性，并概述了实验设计和解释的建议。通过对线粒体生理学研究常用的方法有一个总体的了解，实验者可以开始帮助我们理解这种细胞器，以及它如何影响其他生理表型。