Dual-emissive carbonized polymer dots for the ratiometric fluorescence imaging of singlet oxygen in living cells.

Singlet oxygen (1O2) is a type of reactive oxygen species (ROS), playing a vital role in the physiological and pathophysiological processes. Specific probes for monitoring intracellular 1O2 still remain challenging. In this study, we develop a ratiometric fluorescent probe for the real-time intracellular detection of 1O2 using o-phenylenediamine-derived carbonized polymer dots (o-PD CPDs). The o-PD CPDs possessing dual-excitation-emission properties (blue and yellow fluorescence) were successfully synthesized in a two-phase system (water/acetonitrile) using an ionic liquid tetrabutylammonium hexafluorophosphate as a supporting electrolyte through the electrolysis of o-PD. The o-PD CPDs can act as a photosensitizer to produce 1O2 upon white LED irradiation, in turn, the generated 1O2 selectively quenches the yellow emission of the o-PD CPDs. This quenching behavior is ascribed to the specific cycloaddition reaction between 1O2 and alkene groups in the polymer scaffolds on o-PD CPDs. The interior carbon core can be a reliable internal standard since its blue fluorescence intensity remains unchanged in the presence of 1O2. The ratiometric response of o-PD CPDs is selective toward 1O2 against other ROS species. The developed o-PD CPDs have been successfully applied to monitor the 1O2 level in the intracellular environment. Furthermore, in the inflammatory neutrophil cell model, o-PD CPDs can also detect the 1O2 and other ROS species such as hypochlorous acid after phorbol 12-myristate 13-acetate (PMA)-induced inflammation. Through the dual-channel fluorescence imaging, the ratiometric response of o-PD CPDs shows great potential for detecting endogenous and stimulating 1O2in vivo.

Evaluation of chemotherapeutic response in living cells using subcellular Organelle‒Selective amphipathic carbon dots.

Monitoring of structural changes in subcellular organelles is critical to evaluate the chemotherapeutic response of cells. However, commercial organelle selective fluorophores are easily photobleached, and thus are unsuitable for real-time and long-term observation. We have developed photostable carbon-dot liposomes (CDsomes)-based fluorophores for organellar and suborganellar imaging to circumvent these issues. The CDs synthesized through a mild pyrolysis/hydrolysis process exhibit amphipathic nature and underwent self-assembly to form liposome-like structures (CDsomes). The controlled hydrophilicity or hydrophobicity-guided preparation of CDsomes are used to selectively and rapidly (<1 min) stain nucleolus, cytoplasm, and membrane. In addition, the CDsomes offer universal high-contrast staining not only in fixed cells but also in living cells, allowing real-time observation and morphological identification in the specimen. The as-prepared CDsomes exhibit excitation-dependent fluorescence, and are much more stable under photoirradiation (e.g., ultraviolet light) than traditional subcellular dyes. Interestingly, the CDsomes can be transferred to daughter cells by diluting the particles, enabling multigenerational tracking of suborganelle for up to six generations, without interrupting the staining pattern. Therefore, we believe that the ultra-photostable CDsomes with high biocompatibility, and long-term suborganellar imaging capabilities, hold a great potential for screening and evaluating therapeutic performance of various chemotherapeutic drugs.