High-Resolution Respirometry to assess Bioenergetics in Cells and Tissues using Chamber- and Plate-Based Respirometers

High-resolution respirometry (HRR) allows monitoring oxidative phosphorylation in real-time for analysis of individual cellular energy states and assessment of respiratory complexes using diversified substrate-uncoupler-inhibitor titration (SUIT) protocols. Here, the usage of two high-resolution respirometry devices is demonstrated, and a basic collection of protocols applicable for the analysis of cultured cells, skeletal and heart muscle fibers, and soft tissues such as the brain and liver are presented. Protocols for cultured cells and tissues are provided for a chamber-based respirometer and cultured cells for a microplate-based respirometer, both encompassing standard respiration protocols. For comparative purposes, CRISPR-engineered HEK293 cells deficient in mitochondrial translation causing multiple respiratory system deficiency are used with both devices to demonstrate cellular defects in respiration. Both respirometers allow for comprehensive measurement of cellular respiration with their respective technical merits and suitability dependent on the research question and model under study.

Automated summary

High-resolution respirometry allows real-time monitoring of oxidative phosphorylation and assessing respiratory complexes using diversified SUIT protocols. This study demonstrates the usage of two devices and provides basic protocols for analyzing various cells and tissues. CRISPR-engineered cells are used to showcase cellular respiration defects, highlighting the comprehensive measurement capabilities of both respirometers.