Cai Ning, Lai Alvin Chi-Keung, Liao Kin, Corridon Peter R, Graves David J, Chan Vincent
Wuhan Institute of Technology, School of Chemical Engineering and Pharmacy, Wuhan 430073, China.
Department of Architecture and Civil Engineering, City University of Hong Kong, Tat Chee Avenue, Kowloon Tong, Hong Kong 999077, China.
Polymers (Basel). 2022 May 7;14(9):1913. doi: 10.3390/polym14091913.
Among the new molecular tools available to scientists and engineers, some of the most useful include fluorescently tagged biomolecules. Tools, such as green fluorescence protein (GFP), have been applied to perform semi-quantitative studies on biological signal transduction and cellular structural dynamics involved in the physiology of healthy and disease states. Such studies focus on drug pharmacokinetics, receptor-mediated endocytosis, nuclear mechanobiology, viral infections, and cancer metastasis. In 1976, fluorescence recovery after photobleaching (FRAP), which involves the monitoring of fluorescence emission recovery within a photobleached spot, was developed. FRAP allowed investigators to probe two-dimensional (2D) diffusion of fluorescently-labelled biomolecules. Since then, FRAP has been refined through the advancements of optics, charged-coupled-device (CCD) cameras, confocal microscopes, and molecular probes. FRAP is now a highly quantitative tool used for transport and kinetic studies in the cytosol, organelles, and membrane of a cell. In this work, the authors intend to provide a review of recent advances in FRAP. The authors include epifluorescence spot FRAP, total internal reflection (TIR)/FRAP, and confocal microscope-based FRAP. The underlying mathematical models are also described. Finally, our understanding of coupled transport and kinetics as determined by FRAP will be discussed and the potential for future advances suggested.
在科学家和工程师可用的新型分子工具中，一些最有用的工具包括荧光标记的生物分子。诸如绿色荧光蛋白（GFP）之类的工具已被用于对健康和疾病状态生理学中涉及的生物信号转导和细胞结构动力学进行半定量研究。此类研究聚焦于药物药代动力学、受体介导的内吞作用、核力学生物学、病毒感染和癌症转移。1976年，光漂白后荧光恢复（FRAP）技术得以开发，该技术涉及监测光漂白斑点内的荧光发射恢复情况。FRAP使研究人员能够探究荧光标记生物分子的二维（2D）扩散。从那时起，随着光学、电荷耦合器件（CCD）相机、共聚焦显微镜和分子探针的发展，FRAP技术不断完善。如今，FRAP是一种用于细胞溶质、细胞器和细胞膜运输及动力学研究的高度定量工具。在这项工作中，作者旨在对FRAP的最新进展进行综述。作者介绍了落射荧光斑点FRAP、全内反射（TIR）/FRAP以及基于共聚焦显微镜的FRAP。还描述了其 underlying 数学模型。最后，将讨论我们对由FRAP确定的耦合运输和动力学的理解，并提出未来进展的潜力。 （注：原文中“underlying”未翻译完整，推测可能是“基础的”意思，但按要求不能添加解释，所以保留原文）
