Cell-surface protein-protein interaction analysis with time-resolved FRET and snap-tag technologies.
Cell-surface protein-protein interaction analysis with time-resolved FRET and snap-tag technologies.
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使用时间分辨 FRET 和 snap-tag 技术进行细胞表面蛋白-蛋白相互作用分析。
DOI：
10.1007/978-1-62703-604-7_11
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发表时间：
2013
期刊：
影响因子：
--
通讯作者：
Feinstein,TimothyN
中科院分区：
文献类型：
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作者：
Feinstein,TimothyN
福斯特共振能量转移 (FRET) 是一种邻近依赖性量子效应，可以测量传统形式的荧光或电子显微镜无法看到的蛋白质相互作用和构象变化。然而，FRET 实验通常难以检测瞬时和局部的相互作用，或者在大背景下以低丰度发生的相互作用。该协议描述了一种通过使用时间特异性检测来从串扰激发和所有其他非特异性荧光背景源中分离 FRET 介导的受体发射来提高 FRET 实验的灵敏度和可量化性的方法。
Förster resonance energy transfer (FRET) is a proximity-dependent quantum effect that allows the measurement of protein interactions and conformational changes which are invisible to traditional forms of fluorescence or electron microscopy. However, FRET experiments often have difficulty detecting interactions that are transient and localized or occur in low abundance against a large background. This protocol describes a method of improving on the sensitivity and quantifiability of FRET experiments by using time-specific detection to isolate FRET-mediated acceptor emission from cross-talk excitation and all other sources of nonspecific fluorescence background.