The Research Of Depth Of Field And Autofocus Method In Fluorescent Microscope

As one of the most important optical instruments, the appearance of microscope marks the approaching of atomic era. According to the different imaging principle, microscope can be divided into two types: optical microscope and electron microscope. Fluorescence microscope, one kind of optical microscope and consisted by high-pressure mercury lamp, color filters and optical system, is the basic instrument for immunofluorescent cytochemical technique.With the emerging of CCD and development of computer science and digital image processing, digital microscope system has become the hot spot in the world from 1990 and is wildly used in special microscopes such as fluorescence microscope. Compared with traditional visual microscope, digital microscope is more effective, fast and accurate.Based on fluorescence microscope with visual and digital imaging functions, firstly, the paper pointed out the limitations of traditional methods for calculating depth of field (DOF). Secondly, based on the relationship between DOF and resolution, the paper put up with new calculating methods and the results were more objective and accurate. Finally, according to the small DOF and specialist of object, the paper proposed new evaluating function and searching algorithm, which could be applied in fluorescence microscope. Based on auto-focusing method, the paper accomplished image fusion and obtained fluorescence images with more information and high contrast.The main work is finished in the paper as follows:1 The paper introduced the fluorescence microscope and pointed out the limitations of traditional calculating methods and auto-focusing methods in microscope system.2 The paper proposed new methods for calculating DOF in visual microscope and pointed out it was unnecessary to take account of the visual accommodation in new method.3 The paper proposed new methods for calculating DOF in digital microscope and the results were more objective and accurate.4 The paper put up with new evaluating function and researching algorithm and accomplished the image fusion, which makes the fluorescent images with more information and high contrast.