在科学家和工程师可用的新分子工具中,一些最有用的包括荧光标记的生物分子。工具,如绿色荧光蛋白(GFP),已用于对健康和疾病状态的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。这些研究集中在药物药代动力学,受体介导的内吞作用,核机械生物学,病毒感染,和癌症转移。1976年,光漂白后的荧光恢复(FRAP),这涉及到监测光漂白斑点内的荧光发射恢复,已开发。FRAP允许研究人员探测荧光标记的生物分子的二维(2D)扩散。从那以后,FRAP通过光学的进步得到了完善,电荷耦合器件(CCD)摄像机,共聚焦显微镜,和分子探针。FRAP现在是一种高度定量的工具,用于胞质溶胶中的运输和动力学研究,细胞器,和细胞膜。在这项工作中,作者打算对FRAP的最新进展进行综述.作者包括落射荧光点FRAP,全内反射(TIR)/FRAP,和基于共聚焦显微镜的FRAP。还描述了基本的数学模型。最后,将讨论我们对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。
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.