Real-time temperature monitoring inside living organisms provides a direct measure of their biological activities. However, it is challenging to reduce the size of biocompatible thermometers down to submicrometers, despite their potential applications for the thermal imaging of subtissue structures with single-cell resolution. Here, using quantum nanothermometers based on optically accessible electron spins in nanodiamonds, we demonstrate in vivo real-time temperature monitoring inside Caenorhabditis elegans worms. We developed a microscope system that integrates a quick-docking sample chamber, particle tracking, and an error correction filter for temperature monitoring of mobile nanodiamonds inside live adult worms with a precision of ±0.22°C. With this system, we determined temperature increases based on the worms’ thermogenic responses during the chemical stimuli of mitochondrial uncouplers. Our technique demonstrates the submicrometer localization of temperature information in living animals and direct identification of their pharmacological thermogenesis, which may allow for quantification of their biological activities based on temperature.
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