TIME RESOLVED IMAGING MICROSCOPY - PHOSPHORESCENCE AND DELAYED FLUORESCENCE IMAGING
TIME RESOLVED IMAGING MICROSCOPY - PHOSPHORESCENCE AND DELAYED FLUORESCENCE IMAGING
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DOI：
10.1016/s0006-3495(91)82175-0
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发表时间：
1991-12-01
影响因子：
3.4
通讯作者：
JOVIN, TM
中科院分区：
文献类型：
--
作者：
MARRIOTT, G;CLEGG, RM;JOVIN, TM
研制了一种能够测量时间分辨发光（磷光和延迟荧光）图像的光学显微镜。该技术采用两个锁相机械斩波器和一个慢扫描科学CCD相机连接到一个正常的荧光显微镜。样品被周期性的光脉冲序列照射，并且在每个激发周期结束后的限定时间间隔内记录图像。时间分辨率完全区别于光散射，反射，自发荧光，和外来提示荧光，这通常会降低正常荧光显微镜测量的对比度。时间分辨图像显微镜产生高对比度的图像和特定的结构，可以通过显示一个新的参数，磷光荧光的比率来强调。发光衰减率不同的物体很容易分辨。通过分析一系列不同的图像，可以在显微镜图像的每个像素处测量长寿命发光的寿命，所述图像具有可变的时间延迟。发光衰减率的分布直接显示为图像。几个例子证明了该仪器的实用性和它提供给传统荧光显微镜的互补性。
An optical microscope capable of measuring time resolved luminescence (phosphorescence and delayed fluorescence) images has been developed. The technique employs two phase-locked mechanical choppers and a slow-scan scientific CCD camera attached to a normal fluorescence microscope. The sample is illuminated by a periodic train of light pulses and the image is recorded within a defined time interval after the end of each excitation period. The time resolution discriminates completely against light scattering, reflection, autofluorescence, and extraneous prompt fluorescence, which ordinarily decrease contrast in normal fluorescence microscopy measurements. Time resolved image microscopy produces a high contrast image and particular structures can be emphasized by displaying a new parameter, the ratio of the phosphorescence to fluorescence. Objects differing in luminescence decay rates are easily resolved. The lifetime of the long lived luminescence can be measured at each pixel of the microscope image by analyzing a series of images that differ by a variable time delay. The distribution of luminescence decay rates is displayed directly as an image. Several examples demonstrate the utility of the instrument and the complementarity it offers to conventional fluorescence microscopy.