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Assessments of mitochondrial respiration and mitochondrial content are common in skeletal muscle research and exercise science. However, many sources of technical and biological variation render these analyses susceptible to error. This study aimed to better quantify the reliability of different experimental designs and/or techniques so as to assist researchers to obtain more reliable data.

We examined the repeatability of maximal mitochondrial oxidative phosphorylation in permeabilized muscle fibres via high‐resolution respirometry, and citrate synthase activity (a biomarker for mitochondrial content) in a microplate with spectrophotometery.

For mitochondrial respiration using permeabilized skeletal muscle fibres, the variability was reduced using three chambers and removing outliers compared to two chambers (CV reduced from 12.7% to 11.0%), and the minimal change that can be …