FRET-enabled optical modulation for high sensitivity fluorescence imaging.
FRET-enabled optical modulation for high sensitivity fluorescence imaging.
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DOI：
10.1021/ja100175r
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发表时间：
2010-05-12
影响因子：
15
通讯作者：
Dickson, Robert M.
中科院分区：
文献类型：
--
作者：
Richards, Chris I.;Hsiang, Jung-Cheng;Khalil, Andrew M.;Hull, Nathan P.;Dickson, Robert M.
利用荧光共振能量转移来设计供体电子物理学，以便于信号放大和从高背景中选择性地恢复荧光。这是广义的，使得许多不同的荧光团可以用于光学调制方案，以大大提高荧光成像灵敏度。受体的动态、同时、直接激发使更高能量的供体发射变亮并进行光学调制。外部施加的调制波形使得能够通过解调进行选择性供体荧光提取。通过引入具有显着的、光谱位移的暗态粒子数的受体，所需的激发强度非常低，并且与模拟的增强非常一致。增强与调制频率直接产生暗态寿命在一个简单的系综测量。使用长寿命的Cy5暗态结合Cy3供体，我们证明了从大背景中提取图像，通过同步放大荧光图像恢复（SAFIRe）获得10倍的灵敏度提高。
Fluorescence resonance energy transfer is utilized to engineer donor photophysics for facile signal amplification and selective fluorescence recovery from high background. This is generalized such that many different fluorophores can be used in optical modulation schemes to drastically improve fluorescence imaging sensitivity. Dynamic, simultaneous, direct excitation of the acceptor brightens and optically modulates higher energy donor emission. The externally-imposed modulation waveform enables selective donor fluorescence extraction through demodulation. By incorporating an acceptor with significant, spectrally-shifted, dark state population, necessary excitation intensities are quite low and agree well with simulated enhancements. Enhancement vs. modulation frequency directly yields dark state lifetimes in a simple ensemble measurement. Using the long-lived Cy5 dark state in conjunction with Cy3 donors, we demonstrate image extraction from a large background to yield ≫10-fold sensitivity improvements through Synchronously Amplified Fluorescence Image Recovery (SAFIRe).
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