线粒体是参与真核生物各种代谢过程的重要细胞器。与线粒体相关的细胞死亡机制的成像、靶向和研究引起了人们的极大兴趣。小分子荧光探针已被证明是利用光推进线粒体生物学研究的强大工具。在这项研究中，我们提出了用于这些目的的带有永久正电荷的阳离子尼罗蓝探针的合理设计。阳离子尼罗蓝探针表现出优异的线粒体通透性、独特的溶剂化显色性和抗氧化性。我们观察到与亲脂性溶剂相比，水溶液中的荧光较弱，从而最大限度地减少了细胞质中的背景荧光。此外，我们在温和条件下实现了阳离子尼罗蓝探针的光氧化还原转换。这使我们首次展示了它们在线粒体单分子定位显微镜中的应用，使我们能够观察线粒体裂变和融合行为。与传统的花青荧光团相比，此类染料表现出长时间的抗光漂白性，这可能是由于它们的抗氧化特性。此外，我们将阳离子尼罗蓝探针的应用扩展到紫杉烷类的线粒体特异性递送，促进了药物与细胞器之间直接相互作用的研究。我们在不依赖微管结合的情况下触发细胞死亡的方法为抗癌药物研究和耐药机制提供了宝贵的见解。© 2024 作者。由美国化学会出版。

Mitochondria are essential organelles involved in various metabolic processes in eukaryotes. The imaging, targeting, and investigation of cell death mechanisms related to mitochondria have garnered significant interest. Small-molecule fluorescent probes have proven to be robust tools for utilizing light to advance the study of mitochondrial biology. In this study, we present the rational design of cationic Nile blue probes carrying a permanent positive charge for these purposes. The cationic Nile blue probes exhibit excellent mitochondrial permeability, unique solvatochromism, and resistance to oxidation. We observed weaker fluorescence in aqueous solutions compared to lipophilic solvents, thereby minimizing background fluorescence in the cytoplasm. Additionally, we achieved photoredox switching of the cationic Nile blue probes under mild conditions. This enabled us to demonstrate their application for the first time in single-molecule localization microscopy of mitochondria, allowing us to observe mitochondrial fission and fusion behaviors. Compared to conventional cyanine fluorophores, this class of dyes demonstrated prolonged resistance to photobleaching, likely due to their antioxidation properties. Furthermore, we extended the application of cationic Nile blue probes to the mitochondria-specific delivery of taxanes, facilitating the study of direct interactions between the drug and organelles. Our approach to triggering cell death without reliance on microtubule binding provides valuable insights into anticancer drug research and drug-resistance mechanisms.© 2024 The Authors. Published by American Chemical Society.