Abstract
Characterizing the long-term nanometer-scale interactions between lysosomes and mitochondria in live cells is essential for understanding their functions but remains challenging due to limitations of the existing fluorescent probes. Here, we develop cell-permeable organic fluorescent probes for lysosomes with excellent specificity and high photostability. We also use an existing Atto 647N dye with high brightness and excellent photostability to achieve specific labeling of mitochondria in live cells. Using these probes, we obtain dual-color structured illumination microscopy (SIM) images of dynamic physical lysosome-mitochondrion interactions in live cells at an ~90-nm resolution over a long time course of ~13 min. We successfully record the consecutive dynamic processes of lysosomal fusion and fission, as well as four types of physical lysosome-mitochondrion interactions by super-resolution imaging. Our probes provide an avenue for understanding the functions and the dynamic interplay of lysosomes and mitochondria in live cells.
Highlights
Characterizing the long-term nanometer-scale interactions between lysosomes and mitochondria in live cells is essential for understanding their functions but remains challenging due to limitations of the existing fluorescent probes
We explore the possibility of an existing Atto 647N dye with high fluorescence intensity and excellent photostability as a live-cell mitochondrial marker
We evaluated the cytotoxicity of the lysosomal probes in live cells by using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay
Summary
Characterizing the long-term nanometer-scale interactions between lysosomes and mitochondria in live cells is essential for understanding their functions but remains challenging due to limitations of the existing fluorescent probes. The limitations of existing fluorescent lysosomal and mitochondrial probes, such as photobleaching and a nonspecific background[13,14], hinder the characterization of dynamic physical interactions between lysosomes and mitochondria in live cells. Using our lysosomal probes and Atto 647N, we obtain dual-color SIM images of dynamic physical lysosomea d
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