Ratiometric Covalent Fluorescent Probes for Dynamic Super-Resolution Imaging of Mitochondrial HClO

Area of Science:

• Biomedical Imaging
• Cell Biology
• Molecular Pathogenesis

Background:

• Mitochondrial hypochlorous acid (HClO) dynamics are key to understanding ferroptosis.
• Super-resolution microscopy enables subcellular imaging, but lacks specific mitochondrial HClO probes.
• Existing probes face challenges like off-target labeling, environmental interference, and spectral crosstalk.

Purpose of the Study:

• To develop a novel fluorescent probe for specific, dynamic, super-resolution imaging of mitochondrial HClO.
• To investigate the spatiotemporal relationship between mitochondrial HClO and ferroptosis.
• To overcome limitations of current probes for accurate subcellular HClO detection.

Main Methods:

• Design and screening of novel hypochlorous acid (HClO) fluorescent probes.
• Utilizing a triphenylphosphine and chloroacetyl chloride moiety for specific mitochondrial labeling.
• Employing ratiometric fluorescence and an ultralarge Stokes shift for enhanced detection.
• Performing dynamic super-resolution imaging of mitochondrial HClO during ferroptosis.

Main Results:

• A novel probe, YM-P, was developed for specific, covalent mitochondrial labeling.
• YM-P exhibits ratiometric fluorescence, a low detection limit (35 nM), and minimal spectral crosstalk.
• Dynamic super-resolution imaging revealed mitochondrial cristae alterations precede HClO concentration changes during ferroptosis.
• Observed an initial increase followed by a decrease in mitochondrial HClO during ferroptosis progression.

Conclusions:

• YM-P is a robust tool for dynamic monitoring of mitochondrial HClO in ferroptosis.
• Mitochondrial cristae are more sensitive indicators of ferroptosis onset than HClO concentration.
• The findings provide insights into ferroptosis pathogenesis and support mitochondrial HClO research.