Carbon dot (CD)-based fluorescent probes for rapid and real-time tracking lysosomes movement and ATP monitoring in live cell

The detection of lysosomal dysfunction and abnormal adenosine triphosphate (ATP) levels is crucial due to their roles in various cellular disorders and tumor progression, respectively. In this paper, carbon dot (CD)-based fluorescent probes (CDs-1 and CDs-2) with rapid lysosomal targeting and real-time lysosomal monitoring capability were designed. Through a hydrothermal method, the different nitrogen doping contents of CDs-1 and CDs-2 were controlled by varying the ethylenediamine content while the lysosome-targeting ability derived from their neutral red precursor was preserved. CDs-1 and CDs-2 rapid and effectively targeted lysosomes in both H1975 and 4T1 cells, with Pearson's colocalization coefficients of 0.94 and 0.88 (CDs-1) and 0.84 and 0.84 (CDs-2), closely matching the staining patterns of commercial lysosomal markers. CDs-1 demonstrated high specificity and sensitivity for ATP detection, displaying a linear detection range of 4-44 μM with a detection limit as low as 2.86 μM. CDs-1 enables  monitoring of intracellular ATP level changes in the live cells under etoposide stimulation, while CDs-2 exhibited outstanding photostability and enabled real-time and long-term tracking of lysosomal movement. This study establishes CD-based fluorescent probes as valuable tools for investigating lysosomal function, with potential applications in cell biology research, disease mechanism studies, and drug screening platforms.