Yang Yueli, Bai Xueyang, Hu Fanghao
Department of Chemistry, MOE Key Laboratory of Bioorganic Phosphorus Chemistry and Chemical Biology, Tsinghua University, 100084, Beijing, China.
Nat Commun. 2024 Mar 22;15(1):2578. doi: 10.1038/s41467-024-46904-6.
Optical imaging with photo-controllable probes has greatly advanced biological research. With superb chemical specificity of vibrational spectroscopy, stimulated Raman scattering (SRS) microscopy is particularly promising for super-multiplexed optical imaging with rich chemical information. Functional SRS imaging in response to light has been recently demonstrated, but multiplexed SRS imaging with reversible photocontrol remains unaccomplished. Here, we create a multiplexing palette of photoswitchable polyynes with 16 Raman frequencies by coupling asymmetric diarylethene with super-multiplexed Carbow (Carbow-switch). Through optimization of both electronic and vibrational spectroscopy, Carbow-switch displays excellent photoswitching properties under visible light control and SRS response with large frequency change and signal enhancement. Reversible and spatial-selective multiplexed SRS imaging of different organelles are demonstrated in living cells. We further achieve photo-selective time-lapse imaging of organelle dynamics during oxidative stress and protein phase separation. The development of Carbow-switch for photoswitchable SRS microscopy will open up new avenues to study complex interactions and dynamics in living cells with high spatiotemporal precision and multiplexing capability.
利用光控探针进行光学成像极大地推动了生物学研究。受激拉曼散射（SRS）显微镜具有卓越的振动光谱化学特异性，在提供丰富化学信息的超多重光学成像方面极具潜力。最近已证明了响应光的功能性SRS成像，但具有可逆光控的多重SRS成像仍未实现。在此，我们通过将不对称二芳基乙烯与超多重碳硼烷（碳硼烷开关）偶联，创建了具有16种拉曼频率的光开关聚炔多重调色板。通过对电子光谱和振动光谱的优化，碳硼烷开关在可见光控制下表现出优异的光开关特性，并具有大频率变化和信号增强的SRS响应。在活细胞中展示了不同细胞器的可逆和空间选择性多重SRS成像。我们进一步实现了氧化应激和蛋白质相分离过程中细胞器动态的光选择性延时成像。用于光开关SRS显微镜的碳硼烷开关的开发将为以高时空精度和多重能力研究活细胞中的复杂相互作用和动态开辟新途径。
