A FRAP on TIRF superresolution microscopy system for bioimaging

荧光显微镜的巨大力量最近被表彰，三位策划超分辨率成像的科学家荣获 2014 年诺贝尔化学奖。尽管被誉为活细胞成像的巨大飞跃，但全世界很少有实验室能够真正在活细胞中使用超分辨率。现在还有其他荧光成像模式，可以提高分辨率，还可以实时获取细胞过程动态的定量数据。该项目建议在一台显微镜上结合两种这样的成像模式：全内反射荧光（TIRF）和光漂白后荧光重新分布（FRAP）。具体来说，现有的多激光 TIRF 显微镜将配备必要的硬件，以便能够进行 FRAP 并生成细胞表面蛋白质动态周转的数据。这种新工具对于量化合成世界上最丰富的生物聚合物纤维素的酶复合物以及位于质膜上的膜受体、转运蛋白和其他蛋白质复合物的周转率来说是不可或缺的。

The tremendous power of fluorescence microscopy was recently recognized with the 2014 Nobel Prize in Chemistry to three scientists who have masterminded super resolution imaging. Although hailed as a quantum leap in live cell imaging, there are few labs worldwide who can actually use super resolution in living cells. Other modes of fluorescence imaging are now available that can improve resolution and also obtain quantitative data on the dynamics of cellular processes in real time. This project proposes to combine two such imaging modes, total internal reflectance fluorescence (TIRF) and fluorescence redistribution after photobleaching (FRAP) on one microscope. Specifically, an existing multi-laser TIRF microscope will be retrofitted with the necessary hardware to enable FRAP to be conducted and to generate data on the dynamic turnover of proteins at the cell surface. This new tool will be indispensable for quantifying the turnover of enzyme complexes that synthesize the worlds most abundant biopolymer cellulose, as well as membrane receptors, transporters and other proteins complexes situated at the plasma membrane.