Common methods in mitochondrial research (Review)

Abstract

Mitochondrial abnormalities are primarily seen in morphology, structure and function. They can cause damage to organs, including the heart, brain and muscle, by various mechanisms, such as oxidative stress, abnormal energy metabolism, or genetic mutations. Identifying and detecting pathophysiological alterations in mitochondria is the principal means of studying mitochondrial abnormalities. The present study reviewed methods in mitochondrial research and focused on three aspects: Mitochondrial extraction and purification, morphology and structure and function. In addition to classical methods, such as electron microscopy and mitochondrial membrane potential monitoring, newly developed methods, such as mitochondrial ultrastructural determination, mtdNA mutation assays, metabolomics and analyses of regulatory mechanisms, have also been utilized in recent years. These approaches enable the accurate detection of mitochondrial abnormalities and provide guidance for the diagnosis and treatment of related diseases. Contents1. Introduction 2. Extraction and purification of mitochondria 3. determination of mitochondrial morphology and structure 4. determination of mitochondrial function 5.

Methods to Study Mitochondria: Techniques Used to Study the Effects of Age‐Related Diseases Including Alzheimer's

“…The bioluminescence assay used in our lab is based on light formation from ATP and luciferin catalyzed by luciferase. Light is emitted linearly with respect to ATP concentration and is measured on a luminometer (Reddy, Yin, et al., 2018; Yin & Shen, 2022).…”

“…Electron microscopy has been in use for 50 years and can be used to observe organelles such as mitochondria in eukaryotic cells. Here, it can be used to measure mitochondrial number and length (Granata, Jamnick, & Bishop, 2018; Kandimalla et al., 2018; Manczak et al., 2010, 2018; Reddy, Yin, et al., 2018; Wagner et al., 2003; Yin & Shen, 2022). We have used TEM to observe mitochondrial number, structure, and length in cells and tissue samples (Kandimalla et al., 2018; Manczak et al., 2010, 2018; Reddy, Yin, et al., 2018; Wagner et al., 2003).…”

“…Figure 11 shows graphical representations of data for mitochondrial respiration, maximal respiration, ATP production, and proton leaks obtained using the Seahorse bioanalyzer. All four variables were measured in terms of oxygen consumption rate (OCR), which is used to study oxidative phosphorylation in mitochondria in pmol/min (Yin & Shen, 2022). The results show that mitophagy enhancers (urolithinA, actionin, tomatidine, and nicotinamide riboside) increase mitochondrial respiration in mutantTau‐HT22 cells compared to untransfected HT22 cells.…”

“…Due to differences in excitation wavelengths, JC‐1 produces green fluorescence and TMRE produces red‐orange fluorescence. JC‐1 has higher cytotoxicity compared to TMRE, which can be used for quantitative analysis of MMP due to its low cytotoxicity (Yin & Shen, 2022). ATP production can be used to assess mitochondrial function in cells using bioluminescence assays along with other techniques such as chromatography.…”

Application of fluorescent potential-sensitive probes for the study of living blood and marrow cells

“…Hence, the membrane potential is a universal index of energy conjugation systems [5]. Disturbance of the functional state of MX leads to metabolic disorders in cells [6,7]. Thus, the maintenance of membrane potential serves as an index of the energy balance in MX and the level of metabolic activity of cells [8,9].…”