Fixable Molecular Thermometer for Real-Time Visualization and Quantification of Mitochondrial Temperature

Area of Science:

• Biochemistry
• Cell Biology
• Biomedical Engineering

Background:

• Mitochondrial temperature changes reflect metabolic activity and heat generation.
• Existing fluorescent probes lack stable mitochondrial localization due to dynamic cellular conditions.
• Permanent immobilization is needed for reliable intracellular temperature monitoring.

Purpose of the Study:

• To develop a novel, fixable fluorescent probe for accurate mitochondrial temperature measurement.
• To overcome the limitation of unstable probe localization in dynamic mitochondria.
• To establish a method for visualizing and quantifying mitochondrial temperature in live cells.

Main Methods:

• Design and synthesis of Mito-TEM, a rhodamine B-based fluorescent probe with an anchoring benzaldehyde unit.
• Utilizing the positive charge of the fluorophore for mitochondrial targeting and rotatable substituents for temperature sensitivity.
• Demonstrating probe immobilization via colocalization imaging and applying it to live cell studies.

Main Results:

• Mito-TEM successfully targets and permanently immobilizes within mitochondria.
• The probe exhibits temperature-responsive fluorescence, enabling quantitative measurements.
• Visualized and quantified mitochondrial temperature changes in live cells under various stimuli.

Conclusions:

• Mito-TEM is the first fixable fluorescent molecular thermometer for mitochondria.
• The probe enables stable and accurate monitoring of mitochondrial temperature.
• This technology offers a new tool for studying mitochondrial function and cellular energetics.