Rationale: Measures of mitochondrial respiratory function.
Instrument: Oxygraph-2K (Oroboros Instruments).
Measures: Measures of respiratory activity (oxidative phosphorylation in mitochondria) of biological samples and quantitative data are obtained.
Data Inference: Oxygen concentration and flux.
Controls: Care is taken to maintain the linearity, stability and sensitivity of the signal from polarographic oxygen sensors.
Rationale: Measures of mitochondrial function in primary cell cultures.
Instrument: XF24 Extracellular Flux Analyzer (Seahorse Instrument).
Measures: In vitro oxygen consumption rates (OCR) and extracellular acidification rates (ECAR), indicative of mitochondrial activity and fatty acid oxidation are measured.
Data Inference: Data are dissolved oxygen and pH, which reflect OCR and ECAR.
Control: Care is taken that the biological samples tested and the running buffer is acclimatized to neutral pH at 37oC prior to running the assay.
Rationale: Measures of ATP production, membrane potential and ROS release in isolated mitochondria.
Instrument: Perkin Elmer Victor X Multimode Plate Reader.
Measures: Absorbance, fluorescence and UV capability are measured using multiple-assay technology.
Data Inference: Data are measurements of mitochondrial ATP production rate, reactive oxygen species production rate, and mitochondrial membrane potential.
Control: Care is taken to control and monitor temperature as necessary for assay conditions.