Lee Dong-Ryoung, Kim Young-Duk, Gweon Dae-Gab, Yoo Hongki
Nano Opto-Mechatronics Laboratory, Department of Mechanical Engineering, Korea Advanced Institute of Science and Technology (KAIST), 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, South Korea.
Opt Express. 2013 Jul 29;21(15):17839-48. doi: 10.1364/OE.21.017839.
We propose a new method for high-speed, three-dimensional (3-D) fluorescence imaging, which we refer to as dual-detection confocal fluorescence microscopy (DDCFM). In contrast to conventional beam-scanning confocal fluorescence microscopy, where the focal spot must be scanned either optically or mechanically over a sample volume to reconstruct a 3-D image, DDCFM can obtain the depth of a fluorescent emitter without depth scanning. DDCFM comprises two photodetectors, each with a pinhole of different size, in the confocal detection system. Axial information on fluorescent emitters can be measured by the axial response curve through the ratio of intensity signals. DDCFM can rapidly acquire a 3-D fluorescent image from a single two-dimensional scan with less phototoxicity and photobleaching than confocal fluorescence microscopy because no mechanical depth scans are needed. We demonstrated the feasibility of the proposed method by phantom studies.
我们提出了一种用于高速三维（3-D）荧光成像的新方法，我们将其称为双检测共聚焦荧光显微镜（DDCFM）。与传统的光束扫描共聚焦荧光显微镜不同，在传统方法中，焦斑必须在样品体积上进行光学或机械扫描以重建三维图像，而DDCFM无需深度扫描即可获得荧光发射体的深度。DDCFM在共聚焦检测系统中包括两个光电探测器，每个探测器都有不同尺寸的针孔。通过强度信号的比率，利用轴向响应曲线可以测量荧光发射体的轴向信息。由于无需机械深度扫描，DDCFM可以通过单次二维扫描快速获取三维荧光图像，且与共聚焦荧光显微镜相比，具有更低的光毒性和光漂白效应。我们通过模型研究证明了该方法的可行性。
