对树脂包埋的pHuji进行化学再激活可添加红色荧光，以便与增强绿色荧光蛋白（EGFP）进行同步双色成像。
Chemical reactivation of resin-embedded pHuji adds red for simultaneous two-color imaging with EGFP.
作者信息
Guo Wenyan, Liu Xiuli, Liu Yurong, Gang Yadong, He Xiaobin, Jia Yao, Yin Fangfang, Li Pei, Huang Fei, Zhou Hongfu, Wang Xiaojun, Gong Hui, Luo Qingming, Xu Fuqiang, Zeng Shaoqun
机构信息
Britton Chance Center for Biomedical Photonics, Wuhan National Laboratory for Optoelectronics-Huazhong University of Science and Technology, 1037 Luoyu Road, Wuhan 430074, China.
Department of Biomedical Engineering, Key Laboratory for Biomedical Photonics of Ministry of Education, Huazhong University of Science and Technology, 1037 Luoyu Road, Wuhan 430074, China.
出版信息
Biomed Opt Express. 2017 Jun 15;8(7):3281-3288. doi: 10.1364/BOE.8.003281. eCollection 2017 Jul 1.
The pH-sensitive fluorescent proteins enabling chemical reactivation in resin are useful tools for fluorescence microimaging. EGFP or EYFP is good for such applications. For simultaneous two-color imaging, a suitable red fluorescent protein is an urgent need. Here a pH-sensitive red fluorescent protein, pHuji, is selected and verified to remain pH-sensitive in HM20 resin. We observe 183% fluorescence intensity of pHuji in resin-embeded mouse brain and 29.08-fold fluorescence intensity of reactivated pHuji compared to the quenched state. pHuji and EGFP can be quenched and chemically reactivated simultaneously in resin, thus enabling simultaneous two-color micro-optical sectioning tomography of resin-embedded mouse brain. This method may greatly facilitate the visualization of neuronal morphology and neural circuits to promote understanding of the structure and function of the brain.
能够在树脂中实现化学再激活的pH敏感荧光蛋白是荧光显微成像的有用工具。增强型绿色荧光蛋白（EGFP）或增强型黄色荧光蛋白（EYFP）适用于此类应用。对于同步双色成像，迫切需要一种合适的红色荧光蛋白。在此，我们选择了一种pH敏感的红色荧光蛋白pHuji，并验证其在HM20树脂中仍保持pH敏感性。我们观察到，在树脂包埋的小鼠大脑中，pHuji的荧光强度为原来的183%，再激活后的pHuji荧光强度是淬灭状态下的29.08倍。pHuji和EGFP可以在树脂中同时被淬灭和化学再激活，从而实现对树脂包埋小鼠大脑的同步双色微光学切片断层成像。该方法可能极大地促进神经元形态和神经回路的可视化，以增进对大脑结构和功能的理解。
Zotero 插件