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首页> 外文会议>Conference on Laser Applications to Chemical and Environmental Analysis, Feb 11-13, 2000, Santa Fe, New Mexico >Fluorescence Lifetime Imaging Microscopy (FLIM) on the Single Molecule Level
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Fluorescence Lifetime Imaging Microscopy (FLIM) on the Single Molecule Level

机译：单分子水平的荧光寿命成像显微镜（FLIM）

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摘要

During the last years, several methods for the detection of individual fluorescently labeled molecules have been developed. Especially, for the study of inhomogeneous complex systems single molecule spectroscopy can provide information that is difficult to obtain from ensemble measurements. Due to the limited observation time of individual freely diffusing molecules in solution fluorescence scanning systems including single molecule search programs have been developed to monitor the dynamical behavior of individual immobilized probe molecules on surfaces (for review). Besides polarization measurements, so far only the spectral characteristics, i.e. different emission maxima of different dyes, have been used to detect, identify, and monitor the dynamics of individual fluorescent molecules on surfaces. Therefore we decided to further extend the number of obtainable parameters by developing a scanning system for time-correlated single-photon counting (TCSPC). As shown in Fig. 1 the system consists essentially of an inverted confocal fluorescence microscope and a motion driven controller x,y-microscope stage (SCAN 100x100 and MC2000; Maerzhaeuser, Germany). As excitation source we used a pulsed diode laser emitting at 640 nm with a repetition rate of 50 MHz. The laser diode was driven by an external pulse generator.
机译：在最近几年中，已经开发了几种检测单个荧光标记分子的方法。特别是，对于非均质复杂系统的研究，单分子光谱法可以提供难以从整体测量中获得的信息。由于在溶液中单个自由扩散分子的观察时间有限，因此已开发出包括单分子搜索程序在内的荧光扫描系统，以监测单个固定探针分子在表面上的动力学行为（以供审查）。除偏振测量外，到目前为止，仅光谱特性，即不同染料的不同发射最大值，已被用于检测，识别和监测表面上单个荧光分子的动力学。因此，我们决定通过开发用于时间相关的单光子计数（TCSPC）的扫描系统来进一步扩展可获得参数的数量。如图1所示，该系统主要由倒置共聚焦荧光显微镜和运动驱动控制器x，y显微镜载物台（SCAN 100x100和MC2000；德国Maerzhaeuser）组成。作为激发源，我们使用了以640 nm发射，重复频率为50 MHz的脉冲二极管激光器。激光二极管由外部脉冲发生器驱动。

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