Author Affiliations
Abstract
1 MOE Key Laboratory of Laser Life Science and College of Biophotonics, South China Normal University, Guangzhou 510631, P. R. China
2 Department of Pain Management, the First A±liated Hospital of Jinan University, Guangzhou 510630, P. R. China
Exact interaction mechanism between Bax and Bcl-XL, two key Bcl-2 family proteins, is an interesting and controversial issue. Partial acceptor photobleaching-based quantitative fluorescence resonance energy transfer (FRET) measurement, PbFRET, is a widely used FRET quantification method in living cells. In this report, we implemented pixel-to-pixel PbFRET imaging on a wide-field microscope to map the FRET e±ciency (ET images of single living HepG2 cells co-expressing CFP-Bax and YFP-Bcl-XL. The E value between CFP-Bax and YFP-Bcl-XL was 4.59% in cytosol and 11.31% on mitochondria, conclusively indicating the direct interaction of the two proteins, and the interaction of the two proteins was strong on mitochondria and modest in cytosol.
Bax Bcl-XL protein–protein interaction FRET imaging living cells Journal of Innovative Optical Health Sciences
2020, 13(3): 2050011
华南师范大学生物光子学研究院激光生命科学研究所暨激光生命科学教育部重点实验室, 广东 广州 510631
本文发展了一种针对于固定FRET质粒的单波长激发的E-FRET方法(SDW-E-FRET)。相比于E-FRET方法, SDW-E-FRET只需要测量供体激发时供体通道的荧光强度(IDD)和FRET通道的荧光强度(IDA), 因此该方法可以实现活细胞的快速定量FRET成像。结合双通道荧光显微成像系统, 该方法无需任何机械切换, 定量FRET成像的速度只取决于CCD相机的成像速度, 因而特别适合活细胞实时动态定量FRET成像。在本研究小组发展的双通道荧光显微镜平台上, 应用SDW-E-FRET方法测量了C5V和C17V的FRET效率, 得到了与其它方法测量的一致的结果。
定量FRET成像 FRET探针 单波长激发 实时测量 活细胞 quantitative FRET imaging FRET probe single wavelength excitation real-time measurement living cells
