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This is a In Vitro Study study.

August 19, 2025Open Access

Real-Time Monitoring of the Dynamics and Heterogeneity for Mitochondria and Nucleoli with a Superphotostable Fluorescent Probe by Fluorescence Lifetime Microscopy

QXQianming Xu YWYanxin WangQingdao University YZYulei Zhu

Key Points

The fluorescent probe selectively visualizes mitochondria and nucleoli dynamics during oxidative stress, showing unique fluorescence lifetimes.
The new approach allows for real-time monitoring with minimal interference from the cellular environment.
Assessment using fluorescence lifetime imaging microscopy enables detailed analysis of morphological changes in these organelles.
This technique may facilitate understanding of various diseases linked to mitochondrial and nucleolar functions.

Abstract

Real-time monitoring of the dynamics and heterogeneity of mitochondria and nucleoli is essential for a comprehensive understanding of cellular homeostasis and pathological mechanisms. However, current approaches mainly suffer from two major limitations: (1) Limited permeability through nuclear pore complexes (NPCs) in live cells and (2) vulnerability of intensity-based detection methods to interference from the cellular microenvironment. To overcome these challenges, a red-emissive fluorescent probe (DSM) with high photostability was developed by strategically tuning the π-conjugation between electron-donating and accepting moieties. Gratifyingly, DSM can selectively accumulate in both mitochondria and nucleoli with a distinguishable average fluorescence lifetime after binding with RNA and protein, which enables their dynamic visualization of morphological changes and interplay during oxidative stress by fluorescence lifetime imaging microscopy (FLIM). This research not only paves a new pathway for real-time imaging the physiology of mitochondria and nucleoli but also inspires to guide the pathological study of related diseases.

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