Study on aberration-free focal-depth-extended dark-field microscopy and non-linear imaging analysis

本研究的第一个目的是开发基于焦深扩展暗场显微镜的实用像差校正技术，适用于光学显微镜和电子显微镜。另外，对非线性成像现象的理论分析是第二个目的，即对观测到的图像进行解释。根据这些目标，第一阶段（2001年）发展了包括非线性成像效应的广义成像理论，这是三维成像理论的延伸。在此理论的基础上，编写了评价无像差观测结果的图像仿真程序。第一阶段同时进行了无像差焦深扩展暗场显微镜的设计和构造。此外，还制作了所需的附加控制硬件和处理程序，这些都包含在本显微镜系统中。在研究的下一阶段（2002年），首先对焦深扩展暗场显微镜下无像差观测的评价进行了研究。结果表明，采用动态空心锥照明和适当的图像处理方法，即使在暗场图像中也可以实现幅相分量的分离。利用现有的显微镜对暗场图像进行评价，认为球差的影响是很容易校正的。然而，对于离焦来说，振幅分量和相位分量的分离是非常困难和严重的。在实践中，它可能只适用于在严格焦点设置下的非常薄的样品。最后，无像差焦深扩展暗场显微镜技术可以应用于具有类似三维成像特性的荧光显微镜和自发射显微镜等显微镜。

The first aim of this study is the development of practical aberration correction techniques based on the focal-depth-extended dark-field microscopy, applicable to both optical and electron microscopes. And also, the theoretical analysis on non-linear imaging phenomena is the second aim, to interpret observed images.According to these aims, the generalized imaging theory including non-linear imaging effects, is developed in the first stage (2001), which is the extension of the three dimensional (3D) imaging theory. Under the basis of this theory, image simulation programs to evaluate the aberration-free observation were then coded. The design and construction of an aberration-free focal-depth-extended dark-field microscope were simultaneously carried out in this first stage. In addition, required additional control hardware and processing programs were also made and these were included in the present microscope system.In the next stage (2002) of the study, the evaluation of the aberration free observation under the bases of the focal-depth-extended dark-field microscopy was first examined. Results show that the separation of amplitude and phase components is possible even for dark-field images by the use of the dynamic hollow-cone illumination and the proper image processing. From the evaluation for dark-field images using the present microscope, it is concluded that the effect of the spherical aberration is easily corrected. The separation of the amplitude and the phase components is, however, very difficult and severe for the defocus. In practice, it may be applicable for only very thin samples under the severe focus setting. Finally, the present techniques for the aberration-free focal-depth-extended dark-field microscope can be applicable to microscopes such as fluorescent and self-emission microscopes possess similar 3D imaging characteristics.