Fluorescence lifetime imaging by time-correlated single-photon counting
Fluorescence lifetime imaging by time-correlated single-photon counting
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DOI：
10.1002/jemt.10421
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发表时间：
2004-01-01
影响因子：
2.5
通讯作者：
Biskup, C
中科院分区：
文献类型：
--
作者：
Becker, W;Bergmann, A;Biskup, C
我们提出了一种时间相关单光子计数（TCPSC）技术，该技术允许时间分辨多波长成像与激光扫描显微镜和脉冲激发源相结合。该技术基于四维直方图过程，记录荧光衰减时间内的光子密度、扫描区域的x-y坐标和波长。直方图过程避免了任何时间门控或波长扫描，因此产生了近乎完美的计数效率。时间分辨率仅受探测器的传输时间扩展的限制。该技术可用于几乎任何共聚焦或双光子激光扫描显微镜，并在任何扫描速率下工作。我们演示了用几种染料染色的样品和CFP-YFP FRET的应用。
We present a time-correlated single photon counting (TCPSC) technique that allows time-resolved multi-wavelength imaging in conjunction with a laser scanning microscope and a pulsed excitation source. The technique is based on a four-dimensional histogramming process that records the photon density over the time of the fluorescence decay, the x-y coordinates of the scanning area, and the wavelength. The histogramming process avoids any time gating or wavelength scanning and, therefore, yields a near-perfect counting efficiency. The time resolution is limited only by the transit time spread of the detector. The technique can be used with almost any confocal or two-photon laser scanning microscope and works at any scanning rate. We demonstrate the application to samples stained with several dyes and to CFP-YFP FRET.