Liu Tianyan, Kompa Julian, Ling Jing, Lardon Nicolas, Zhang Yuan, Chen Jingting, Reymond Luc, Chen Peng, Tran Mai, Yang Zhongtian, Zhang Haolin, Liu Yitong, Pitsch Stefan, Zou Peng, Wang Lu, Johnsson Kai, Chen Zhixing
College of Future Technology, Institute of Molecular Medicine, National Biomedical Imaging Center, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, Peking University, Beijing 100871, China.
Peking-Tsinghua Center for Life Science, Academy for Advanced Interdisciplinary Studies, State Key Laboratory of Membrane Biology, Peking University, Beijing 100871, China.
ACS Cent Sci. 2024 Oct 2;10(10):1933-1944. doi: 10.1021/acscentsci.4c00616. eCollection 2024 Oct 23.
Rhodamines have been continuously optimized in brightness, biocompatibility, and color to fulfill the demands of modern bioimaging. However, the problem of phototoxicity caused by the excited fluorophore under long-term illumination has been largely neglected, hampering their use in time-lapse imaging. Here we introduce cyclooctatetraene (COT) conjugated rhodamines that span the visible spectrum and exhibit significantly reduced phototoxicity. We identified a general strategy for the generation of Gentle Rhodamines, which preserved their outstanding spectroscopic properties and cell permeability while showing an efficient reduction of singlet-oxygen formation and diminished cellular photodamage. Paradoxically, their photobleaching kinetics do not go hand in hand with reduced phototoxicity. By combining COT-conjugated spirocyclization motifs with targeting moieties, these Gentle Rhodamines compose a toolkit for time-lapse imaging of mitochondria, DNA, and actin, and synergize with covalent and exchangeable HaloTag labeling of cellular proteins with less photodamage than their commonly used precursors. Taken together, the Gentle Rhodamines generally offer alleviated phototoxicity and allow advanced video recording applications, including voltage imaging.
罗丹明类化合物在亮度、生物相容性和颜色方面不断得到优化，以满足现代生物成像的需求。然而，长期光照下激发态荧光团引起的光毒性问题在很大程度上被忽视了，这限制了它们在延时成像中的应用。在此，我们介绍了与环辛四烯（COT）共轭的罗丹明类化合物，它们覆盖可见光谱，且光毒性显著降低。我们确定了一种生成温和型罗丹明的通用策略，该策略在保留其出色光谱特性和细胞通透性的同时，能有效减少单线态氧的形成并减轻细胞光损伤。矛盾的是，它们的光漂白动力学与降低的光毒性并不一致。通过将COT共轭的螺环化基序与靶向部分相结合，这些温和型罗丹明构成了一个用于线粒体、DNA和肌动蛋白延时成像的工具包，并与细胞蛋白的共价和可交换卤代标签标记协同作用，相比常用的前体，光损伤更小。综上所述，温和型罗丹明总体上减轻了光毒性，并允许进行包括电压成像在内的高级视频记录应用。
