Non fitting based FRET-FLIM analysis approaches applied to quantify protein-protein interactions in live cells.
Non fitting based FRET-FLIM analysis approaches applied to quantify protein-protein interactions in live cells.
复制标题
DOI：
10.1007/s12551-011-0047-6
复制
发表时间：
2011-06-01
影响因子：
--
通讯作者：
Tramier, Marc
中科院分区：
文献类型：
--
作者：
Padilla-Parra, Sergi;Auduge, Nicolas;Tramier, Marc
在过去的几年中，定量荧光显微镜和纳米显微镜的新成像方法已经开发出来，并开始广泛应用于生物学问题，如蛋白质相互作用的定位和定量。荧光共振能量转移（FRET）检测荧光寿命成像显微镜（FLIM）目前不仅在生物物理学或化学，而且在生物医学中，由于新的技术进步，也在数据处理的新发展。FRET FLIM可以是一个非常有用的工具，以确定蛋白质相互作用发生在单个活细胞。在这篇综述中，我们强调的重要性，提高采集速度时，在体内工作采用时域FLIM。新的基于几何的非拟合方法的开发允许在不需要高计数统计的情况下确定相互作用供体的分数，并且因此允许在FRET-FLIM中执行高速采集仍然是定量的。
New imaging methodologies in quantitative fluorescence microscopy and nanoscopy have been developed in the last few years and are beginning to be extensively applied to biological problems, such as the localization and quantification of protein interactions. Fluorescence resonance energy transfer (FRET) detected by fluorescence lifetime imaging microscopy (FLIM) is currently employed not only in biophysics or chemistry but also in bio-medicine, thanks to new advancements in technology and also new developments in data treatment. FRET-FLIM can be a very useful tool to ascertain protein interactions occurring in single living cells. In this review, we stress the importance of increasing the acquisition speed when working in vivo employing Time-Domain FLIM. The development of the new mathematical-based non-fitting methods allows the determining of the fraction of interacting donor without the requirement of high count statistics, and thus allows the performing of high speed acquisitions in FRET-FLIM to still be quantitative.