Fluorescence lifetime imaging microscopy(FLIM)is increasingly used in biomedicine,material science,chemistry,and other related research fields,because of its advantages of high specificity and sensitivity in monitorin...Fluorescence lifetime imaging microscopy(FLIM)is increasingly used in biomedicine,material science,chemistry,and other related research fields,because of its advantages of high specificity and sensitivity in monitoring cellular microenvironments,studying interaction between proteins,metabolic state,screening drugs and analyzing their efficacy,characterizing novel materials,and diagnosing early cancers.Understandably,there is a large interest in obtaining FLIM data within an acquisition time as short as possible.Consequently,there is currently a technology that advances towards faster and faster FLIM recording.However,the maximum speed of a recording technique is only part of the problerm.The acquisition time of a FLIM image is a complex function of many factors.These include the photon rate that can be obtained from the sample,the amount of information a technique extracts from the decay functions,the fficiency at which it determines fluorescence decay parameters from the recorded photons,the demands for the accuracy of these parameters,the number of pixels,and the lateral and axial resolutions that are obtained in biological materials.Starting from a discussion of the parameters which determine the acquisition time,this review will describe existing and emerging FLIM techniques and data analysis algo-rithms,and analyze their performance and recording speed in biological and biomedical applications.展开更多
传统的时间相关单光子计数(Time-correlated single photon counting, TCSPC)受计数率低、一维测量和采集时间长的限制,难以在生物医学中的推广.评述TCSPC的机理和应用,指出TCSPC是探测周期光信号内的单个光子、测量光子探测时间以及构...传统的时间相关单光子计数(Time-correlated single photon counting, TCSPC)受计数率低、一维测量和采集时间长的限制,难以在生物医学中的推广.评述TCSPC的机理和应用,指出TCSPC是探测周期光信号内的单个光子、测量光子探测时间以及构建信号周期内光子分布与时间关系的一种技术,具有多维光子采集特性,计数率近似当前可用的探测器容量.该技术可采集光子对波长和空间座标的分布,时间尺度在皮秒量级,在实验开始时可进行二次计数.多维TCSPC技术拥有近似于理想的计数效率,其时间分辨率仅受探测器渡越时间的离散限制.故TCSPC在生物医学光谱学中极具应用价值.典型应用包括,时域光学X线断层照相、记录生物系统内的瞬态现象、荧光寿命成像显微术、活细胞内荧光共振能量转移实验,以及用荧光关联谱探索染料-蛋白复合体.展开更多
基金support from the National Key R&D Program of China(2017YFA0700500)National Natural Science Foundation of China(61775144/61525503/61620106016/61835009/81727804)+2 种基金(Key)Project of Department of Education of Guangdong Province(2015KGJHZ002/2016KCXTD007)Guangdong Natural Science Foundation(2014A030312008,2017A030310132,2018A030313362)Shenzhen Basic Research Project(JCYJ20170818144012025/JCYJ20170818141701667/JCYJ20170412105003520/JCYJ20150930104948169).
文摘Fluorescence lifetime imaging microscopy(FLIM)is increasingly used in biomedicine,material science,chemistry,and other related research fields,because of its advantages of high specificity and sensitivity in monitoring cellular microenvironments,studying interaction between proteins,metabolic state,screening drugs and analyzing their efficacy,characterizing novel materials,and diagnosing early cancers.Understandably,there is a large interest in obtaining FLIM data within an acquisition time as short as possible.Consequently,there is currently a technology that advances towards faster and faster FLIM recording.However,the maximum speed of a recording technique is only part of the problerm.The acquisition time of a FLIM image is a complex function of many factors.These include the photon rate that can be obtained from the sample,the amount of information a technique extracts from the decay functions,the fficiency at which it determines fluorescence decay parameters from the recorded photons,the demands for the accuracy of these parameters,the number of pixels,and the lateral and axial resolutions that are obtained in biological materials.Starting from a discussion of the parameters which determine the acquisition time,this review will describe existing and emerging FLIM techniques and data analysis algo-rithms,and analyze their performance and recording speed in biological and biomedical applications.
文摘传统的时间相关单光子计数(Time-correlated single photon counting, TCSPC)受计数率低、一维测量和采集时间长的限制,难以在生物医学中的推广.评述TCSPC的机理和应用,指出TCSPC是探测周期光信号内的单个光子、测量光子探测时间以及构建信号周期内光子分布与时间关系的一种技术,具有多维光子采集特性,计数率近似当前可用的探测器容量.该技术可采集光子对波长和空间座标的分布,时间尺度在皮秒量级,在实验开始时可进行二次计数.多维TCSPC技术拥有近似于理想的计数效率,其时间分辨率仅受探测器渡越时间的离散限制.故TCSPC在生物医学光谱学中极具应用价值.典型应用包括,时域光学X线断层照相、记录生物系统内的瞬态现象、荧光寿命成像显微术、活细胞内荧光共振能量转移实验,以及用荧光关联谱探索染料-蛋白复合体.
