Despite the urgent need to image living specimens for cutting-edge biological research, most existing fluorescent labeling methods suffer from either poor optical properties or complicated operations required to realize cell-permeability and specificity. In this study, we introduce a method to overcome these limits—taking advantage of the intrinsic affinity of bright and photostable fluorophores, no matter if they are supposed to be live-cell incompatible or not. Incubated with living cells and tissues in particular conditions (concentration and temperature), some Atto and BODIPY dyes show live-cell labeling capability for specific organelles without physical cell-penetration or chemical modifications. Notably, by using Atto 647N as a live-cell mitochondrial marker, we obtain 2.5-time enhancement of brightness and photostability compared with the most commonly used SiR dye in long-term imaging. Our strategy has expanded the scientist's toolbox for understanding the dynamics and interactions of subcellular structures in living specimens.

尽管迫切需要对生物标本成像以进行前沿的生物学研究，但大多数现有的荧光标记方法都存在光学性能差或实现细胞通透性和特异性所需的复杂操作。在这项研究中，我们介绍了一种克服这些限制的方法-利用明亮和光稳定的荧光团的固有亲和力，无论它们是否与活细胞不相容。在特定条件下（浓度和温度）与活细胞和组织一起孵育，某些Atto和BODIPY染料显示了对特定细胞器的活细胞标记能力，而无需进行物理细胞渗透或化学修饰。值得注意的是，通过使用Atto 647N作为活细胞线粒体标记，我们得到2。与长期成像中最常用的SiR染料相比，亮度和光稳定性提高了5倍。我们的策略扩大了科学家的工具箱，以了解活体标本中亚细胞结构的动力学和相互作用。