Abstract：
Developing fluorogenic probes for simultaneous live cell labeling of multiple targets is crucial for understanding complex cellular events. The emerging [4+1] cycloaddition between tetrazine and isonitriles holds promise as a bioorthogonal tool, yet existing tetrazine probes lack reactivity and fluorogenicity. Here, we present the development of a series of tetrazine-functionalized bioorthogonal probes. By incorporating pyrazole adducts into the fluorophore scaffolds, the post-reacted probes displayed remarkable fluorescence turn-on ratios, up to 3184-fold. Moreover, these modifications are generalizable to various fluorophores, enabling a broad emission range from 473 to 659 nm. Quantum chemical calculations further elucidate the turn-on mechanisms. These probes enable the simultaneous labeling of multiple targets in live cells, without the need for a washing step. Consequently, our findings pave the way for advanced multiplex imaging and detection techniques for cellular studies.
摘要：
开发用于多个靶标的同时活细胞标记的荧光探针对于理解复杂的细胞事件至关重要。四嗪和异腈之间新兴的[41]环加成有望成为生物正交工具，然而现有的四嗪探针缺乏反应性和荧光性。这里,我们介绍了一系列四嗪功能化的生物正交探针的开发。通过将吡唑加合物掺入荧光团支架中，反应后的探针显示出显著的荧光开启率，高达3184倍。此外，这些修饰可推广到各种荧光团，使从473到659nm的宽发射范围。量子化学计算进一步阐明了开启机制。这些探针能够同时标记活细胞中的多个靶标，不需要洗涤步骤。因此,我们的发现为细胞研究的先进的多重成像和检测技术铺平了道路.
