Introduction
The O2k-Fluorescence LED2-Module is an extension of the OROBOROS Oxygraph-2k for combining high-resolution respirometry and fluorometric measurements. We applied the Fluo-Sensor Green for measurement of H2O2 production with Amplex® Ultrared in mouse heart mitochondria. Experiments are selected to demonstrate the importance of applying ETS-competent substrate states, which are critically evaluated by respiratory performance and considering experimental oxygen levels when measuring mitochondrial H2O2 production.