Electronic Preresonance Stimulated Raman Scattering Imaging of Red-Shifted Proteorhodopsins: Toward Quantitation of the Membrane Potential
Electronic Preresonance Stimulated Raman Scattering Imaging of Red-Shifted Proteorhodopsins: Toward Quantitation of the Membrane Potential
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DOI：
10.1021/acs.jpclett.9b01337
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发表时间：
2019-08-01
影响因子：
5.7
通讯作者：
Cheng, Ji-Xin
中科院分区：
文献类型：
--
作者：
Lee, Hyeon Jeong;Huang, Kai-Chih;Cheng, Ji-Xin
电压成像可以绘制活细胞的膜电位。然而，目前基于强度的成像方法仅限于相对膜电位的变化，缺少膜电压绝对值所传达的重要信息。这一挑战源于影响信号强度的各种因素，如浓度、光照强度和光漂白。在这里，我们展示了电子共振高光谱受激拉曼散射(EPR-hSRS)用于光谱检测膜电压，使用在大肠杆菌中表达的近红外吸收微生物视紫红质。这种新开发的近红外活性微生物视紫红质使电子预共振SRS成像具有高灵敏度。通过光谱图谱，我们在单个大肠杆菌细胞的指纹区识别了电压敏感的SRS峰。这些光谱特征为定量活细胞的绝对膜电压提供了一种新的方法。
Voltage imaging allows mapping of the membrane potential in living cells. Yet, current intensity-based imaging approaches are limited to relative membrane potential changes, missing important information conveyed by the absolute value of the membrane voltage. This challenge arises from various factors affecting the signal intensity, such as concentration, illumination intensity, and photobleaching. Here, we demonstrate electronic preresonance hyperspectral stimulated Raman scattering (EPR-hSRS) for spectroscopic detection of the membrane voltage using a near-infrared-absorbing microbial rhodopsin expressed in E. coli. This newly developed near-infrared active microbial rhodopsin enables electronic preresonance SRS imaging at high sensitivity. By spectral profiling, we identified voltage-sensitive SRS peaks in the fingerprint region in single E. coli cells. These spectral signatures offer a new approach for quantitation of the absolute membrane voltage in living cells.