A scalable plate reader-based protocol to measure mitochondrial respiration in embryonic hearts

Mitochondrial function is central to embryonic development, yet its assessment in the embryonic heart is hampered by limited tissue availability. Here, we present a protocol to measure mitochondrial oxygen consumption rates directly from embryonic mouse hearts using a fluorescence-based plate reader assay. We describe steps for heart isolation, plate setup, and acquisition of signal profiles. We then detail procedures for normalization and rate conversion. This protocol enables real-time readings with a low input requirement of one heart.