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Updated: May 28, 2026
Simultaneous Measurement of Mitochondrial Calcium and Mitochondrial Membrane Potential in Live Cells by Fluorescent Microscopy
Published on: January 24, 2017
A highly sensitive and simultaneous bifunctional fluorescent probe for detecting mitochondrial viscosity and
Manlin Fu1, Jiarong Zhou1, Qingwen Li1
1Institute of Natural Products for Chemical Biology, Key Laboratory of Endangered Plants Sustainable Utilization of Taizhou, School of Pharmaceutical Sciences, Taizhou University, Zhejiang, 318000, PR China.
Background:
Mitochondrial dysfunction and dysregulated enzymatic activity are hallmarks of liver cancer progression. Mitochondrial viscosity reports changes in the organelle microenvironment, whereas nitroreductase (NTR) activity is closely associated with tumor hypoxia and metabolic reprogramming. However, few probes enable reliable, simultaneous readout of both biomarkers in living systems. A robust fluorescent tool for dual-parameter imaging of mitochondrial viscosity and NTR activity would facilitate mechanistic studies and support potential tumor diagnosis.
Results:
A novel naphthalimide-based bifunctional fluorescent probe, NCN-NTR, was designed and synthesized for the highly sensitive and simultaneous detection of mitochondrial viscosity and NTR activity in living systems via dual-emission channels. NCN-NTR displays a significant red fluorescence enhancement at 640 nm under high-viscosity conditions due to restricted intramolecular rotation, and exhibits a distinct 445 nm fluorescence at 390 nm upon NTR-mediated cleavage. The probe shows excellent selectivity, photostability, and pH tolerance, enabling reliable imaging in complex biological systems. Confocal microscopy confirmed its mitochondrial localization and enabled visualization of microenvironmental changes during hypoxia and starvation. Importantly, the probe effectively discriminated cancer cells from normal cells and produced rapid, tumor-associated fluorescence signals in vivo. These results collectively highlight the probe's capacity for sensitive, real-time dual-modal bioimaging.
Significance:
NCN-NTR integrates organelle targeting with dual-emission sensing to simultaneously report mitochondrial viscosity and NTR activity in real time. Its performance in cellular and in vivo models highlights its potential for mechanistic studies of tumor-associated microenvironmental alterations and for precision fluorescence imaging with potential applications in cancer diagnosis.
