Chinese Optics Letters, 2010, 8 (10): 947, Published Online: Oct. 19, 2010
Improving FRET efficiency measurement in confocal microscopy imaging Download: 864次
荧光共振能量转移效率 PFRET算法 E-FRET方程 共聚焦显微镜 光谱串扰 170.1790 Confocal microscopy 100.3008 Image recognition, algorithms and filters 110.2960 Image analysis
Abstract
Spectral bleedthrough (SBT) ratio is dependent on the level of fluorescence intensity in confocal imaging. Precision Forster resonance energy transfer (FRET) algorithm corrects SBT ratio according to fluorescence intensity and avoids over- or under-estimation of SBT ratio. In this letter, we propose a new method to accurately measure the FRET efficiency of FRET plasmid in single living cells by combining the calculation of SBT in precision FRET algorithm with E-FRET formulae. We also use this method to measure the FRET efficiency of FRET-Bid, and find that in healthy A549 cells it is about 15%, which is verified by FRET acceptor photobleaching method.
Huaina Yu, Tongsheng Chen, Junle Qu. Improving FRET efficiency measurement in confocal microscopy imaging[J]. Chinese Optics Letters, 2010, 8(10): 947.