Analysis of mitochondrial oxygen consumption and hydrogen peroxide release from cardiac mitochondria using electrochemical multi-sensors

Mitochondria are the primary sites of oxygen (O2) consumption and energy metabolism in most cell types, but also produce reactive species (ROS) that contribute to a wide array pathological physiological processes. Accordingly, simultaneous monitoring mitochondrial ROS release rate (OCR) from cells preparations is an attractive investigative approach biological research, particularly when sample quantity scarce. This paper presents development sensitive multi-sensor device capable measuring production OCR samples single micro-chamber assay. Sensor sensitivities for O2 hydrogen peroxide (H2O2; major released by mitochondria cells) 4.32 nA/?M 54.89 nA/?M, respectively, with limits detection 2.9 ?M 58.36 nM, respectively. Proof-of-concept studies isolated rat cardiac tissue (5 µg protein) demonstrate expected 3 – 4 fold increase H2O2 over basal following addition respiratory substrates, comparatively small change OCR. The subsequent adenosine diphosphate (ADP) decreased 73% (p < 0.01) increased 168% 0.01), consistent established shifts membrane potential electron flow ADP-limited (State 4) ADP-stimulated 3) state. These validate results use novel simultaneously scarce samples, utility non-destructive integrative study cellular function.