基于受体检测的Förster 共振能量转移的 FLIM 显微镜。
A FLIM Microscopy Based on Acceptor-Detected Förster Resonance Energy Transfer.
机构信息
Department of Chemistry, University of Nebraska - Lincoln, Lincoln, Nebraska 68588-0304, United States.
Tecnologico de Monterrey, School of Engineering and Sciences, Av. Eugenio Garza Sada 2501 Sur, Monterrey, Nuevo Leon 64849, Monterrey, Mexico.
出版信息
Anal Chem. 2021 Mar 23;93(11):4841-4849. doi: 10.1021/acs.analchem.0c04492. Epub 2021 Mar 10.
Time-resolved donor-detected Förster resonance energy transfer (trDDFRET) allows the observation of molecular interactions of dye-labeled biomolecules in the ∼10-100 Å region. However, we can observe longer-range interactions when using time-resolved acceptor-detected FRET (trADFRET), since the signal/noise ratio can be improved when observing the acceptor emission. Therefore, we propose a new methodology based on trADFRET to construct a new fluorescence lifetime microscopy (FLIM-trADFRET) technique to observe biological machinery in the range of 100-300 Å in vivo, the last frontier in biomolecular medicine. The integrated trADFRET signal is extracted in such a way that noise is canceled, and more photons are collected, even though trADFRET and trDDFRET have the same rate of transfer. To assess our new methodology, proof of concept was demonstrated with a set of well-defined DNA scaffolds.
时间分辨供体探测Förster 共振能量转移（trDDFRET）允许观察染料标记生物分子在约 10-100 Å 区域的分子相互作用。然而，当使用时间分辨受体探测 FRET（trADFRET）时，我们可以观察到更长的相互作用，因为当观察受体发射时，可以提高信号/噪声比。因此，我们提出了一种基于 trADFRET 的新方法，构建了一种新的荧光寿命显微镜（FLIM-trADFRET）技术，以观察体内 100-300 Å 的生物机制，这是生物分子医学的最后一个前沿领域。以这种方式提取集成的 trADFRET 信号，即使 trADFRET 和 trDDFRET 具有相同的转移速率，也可以消除噪声并收集更多的光子。为了评估我们的新方法，用一组定义明确的 DNA 支架证明了概念验证。
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