光纤共焦扫描显微成像技术的研究
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摘要
单模光纤激光共焦扫描显微成像系统是研究微观结构的一种新技术，其本质是抑制目标物体离焦面所反射的光线进入系统，从而抑制这些离焦光线造成像点弥散斑增大的影响，使系统的横向与轴向分辨率得到大幅度的提高，能实现亚微米级的层析。这对于研究微观物体的表观及层析结构具有重大意义，在生物、微电子等领域具有重要的应用前景。
本文在透镜成像理论的基础上，系统、深入地分析了共焦扫描显微成像的机理，论述了应用单模光纤的激光共焦扫描显微成像系统的优点；进行了总体方案的论证，并设计确定了单模光纤激光共焦扫描显微成像系统的总体方案；从理论上推导分析了分辨率要求与试验系统中相关器件主要参数之间的关系，分析了系统耦合效率和渐晕现象对光学系统的设计要求；完成了方案中光学系统和二维扫描控制电路的设计，并在电路设计中采用了用软件解决检流计式光学扫描器(振镜)非线性问题的新方法，能够实现较为理想的二维模拟扫描；完成了高增益、低噪声和低失真的探测接收系统的设计和调试。
Single mode optical fiber laser confocal scanning microscopy is a new technology in microstructure studying. The speciality of the system is limiting the out-of-focus light reflected from out-of-focus plane in object to restrict the blur of image. Therefore, the plane resolution and axial resolution of the system could be improved to sub-micron grade. It was important in studying the surface and layers of microstructure, and has bright prospect especially in the applied domain of biology and microelectronics. On the base of studying imaging theory of lens, the imaging theory of laser confocal scanning microscopy was analyzed in detail in this paper, and the advancement of that the optical fiber was applied to the system was described; on the base of completed the demonstration for whole project, the experiment scheme was designed; the relationship between the main parameters of key devices and the resolution was deduced, and the requirements of coupling efficiency and vignetting effects to optical system was analyzed; the design of optical system and the planar scanning controlling circuit was completed; a new method was put forward to resolved the inherent non-liner scanning problem of the galvanometer scanner by using software liner controlling in circuit design, and the perfect planar scanning was realized; at last the low noise, high multiple and non-distortion amplify circuit of photoelectric detector was completed.
本文在透镜成像理论的基础上，系统、深入地分析了共焦扫描显微成像的机理，论述了应用单模光纤的激光共焦扫描显微成像系统的优点；进行了总体方案的论证，并设计确定了单模光纤激光共焦扫描显微成像系统的总体方案；从理论上推导分析了分辨率要求与试验系统中相关器件主要参数之间的关系，分析了系统耦合效率和渐晕现象对光学系统的设计要求；完成了方案中光学系统和二维扫描控制电路的设计，并在电路设计中采用了用软件解决检流计式光学扫描器(振镜)非线性问题的新方法，能够实现较为理想的二维模拟扫描；完成了高增益、低噪声和低失真的探测接收系统的设计和调试。
Single mode optical fiber laser confocal scanning microscopy is a new technology in microstructure studying. The speciality of the system is limiting the out-of-focus light reflected from out-of-focus plane in object to restrict the blur of image. Therefore, the plane resolution and axial resolution of the system could be improved to sub-micron grade. It was important in studying the surface and layers of microstructure, and has bright prospect especially in the applied domain of biology and microelectronics. On the base of studying imaging theory of lens, the imaging theory of laser confocal scanning microscopy was analyzed in detail in this paper, and the advancement of that the optical fiber was applied to the system was described; on the base of completed the demonstration for whole project, the experiment scheme was designed; the relationship between the main parameters of key devices and the resolution was deduced, and the requirements of coupling efficiency and vignetting effects to optical system was analyzed; the design of optical system and the planar scanning controlling circuit was completed; a new method was put forward to resolved the inherent non-liner scanning problem of the galvanometer scanner by using software liner controlling in circuit design, and the perfect planar scanning was realized; at last the low noise, high multiple and non-distortion amplify circuit of photoelectric detector was completed.
引文
1.1 M. Minsky, U. S. patent 3013467, Microscopy Apparatus, Dec. 19, 1961(Field Nov. 7, 1957)
1.2 Stan Reich, THE USE OF ELECTRO-MECHANICAL MIRROR SCANNING DEVICE, SPIE,Vol. 84, 1976, 47-56
1.3 D. K. Hamiliton, T. Wilson, C. J. Sheppard. Experiment observations of depth discrimation properties of scanning microscopes. Optical Letter, Vol. 6, No. 12, 1983, 625-626
1.4 I. J. Cox, Scanning Optical Fluorescence Microscopy, J. Microscopy, Vol.133, 1984, 149-154
1.5 B. Bertero, Analytic inversion formula for confocal scanning microscopy, J. Opt. Soc. Am. A, Vol. 4, No. 9, 1987, 1748-1750
1.6 T. Wilson, A. R. Carline, Three-dimensional imaging in confocal imaging, J. Microscopy, Vol.149, 1987, 61-66
1.7 K. Carlson, Three-dimensional Microscopy using a Confocal Laser Scanning Microscope, Optical Letters, Vol. 10, No. 2, 1985, 53-55
1.8 Knell Carlson, Niles Aslund, Confocal imaging for 3-d digital Microscopy, Applied Optics, Vol. 26, No. 16, 1987, 3232-3238
1.9 David N. Sitter, William T. Rhode, Three-dimension imaging: a space invariant model for space variant system, Applied Optics, Vol. 29, No. 26, 1990, 3789-3794
1.10 张平，陈德，吴震，黄俊，共焦激光扫描荧光显微镜的扫描系统研究，应用光学，Vol．16，No．3，1995，33-38
1.11 刘峰，刘明，陶纯堪，聂守平，李爱明，激光共焦快速扫描显微成像的实现，应用光学，Vol．16，No．5，1995，27-29
1.12 任雅萍，杨玉孝，赵明涛，王林祥，层析式激光三维测量技术，激光与红外，Vol．27，No．4，1997，226-227
1.13 杨利松，王桂英，徐至展，光学层析成像机理分析，激光与红外，Vol．27，No．4，1997，239-241
1.14 富光华，刘新，实时共焦扫描光学显微镜，半导体光电，Vol．13，No．1，1992，
30-35
1． 15 L. Giniunas, R. Juskaitis, SCANNING FIBER-OPTIC MICROSCOPE, Electronics Letters, Vol.27, No. 9, 1991,724-726
1． 16 M. E. Marhic, S. E. Schscham, M. Epstein, Misalignment in imaging multifibers, Applied Optics, Vol.17, No. 21, 1978, 3503-3507
1． 17 T. Wilson. Images of Phase Edges in Conventional and Scanning Optical Microscopes. Applied Optics, Vol.20, No. 18, 1981, 3238-3244
1． 18 Min Gu, X. Gan, C. J. R. Sheppard, Three-dimensional coherent functions in a fiber-optical confocal scanning microscope, J. Opt. Soc. Am. A, Vol.8, No. 7, 1991, 1019-1025
1． 19 Kenneth P. Ghiggion, Martin R.Harris, Paul G. Spizzirri, Fluorescence lifetime measurements using a novel fiber-optic laser scanning confocal microscope, American Institute of Physics, Vol.66, No.5, 1992, 2999-3002
1． 20 N. Cerre, F. de Fornel, J. P. Goudonnet, Reflection scanning microscope: influence of the distance and the orientation between the fiber and the sample, Optical Society of America, Vol. 10, No.9, 1992, 1909-1917
1． 21 R. Juskaitis, N. Rea, Tony Wilson, Fiber-optic based confocal microscopes using laser detection, Optics Communications, Vol.99, No. 1, 2, 1993, 105-113
1． 22 Arthur F. Gmitro, David Aziz, Confocal microscopy through a fiber-optic imaging bundle, Optics Letters, Vol.18, No. 8, 1993, 565-567
1． 23 Peter M. Delaney, Martin R. Harris, Roger G. King, Fiber-optic laser scanning confocal microscope suitable for fluorescence imaging, Applied Optics, Vol.33, No. 4, 1994, 573-577
1． 24 R. Juskaitis, T. Wilson, Direct-view fiber-optic confocal microscope, Optical Society of America, Vol. 19, No. 22, 1994, 1500-1501
1． 25 David Aziz, Arthur F. Gmitro, Imaging performance of the fiber-optic image-bundle confocal microscope, SPIE, Vol. 2083, 1994
1． 26 N. P. Rea, R. Juskaitis, T. Wilson, Semiconductor laser confocal and interference microscope, Optics Communication, Vol. 125, 1996, 158-167
1.27 宋葵，李增发，光学显微镜的最新发展，物理，Vol．22，No．2，1992，81-86
1.28 魏高尚，向际鹰，吴震，张平，单模光纤共焦扫描显微系统研究，应用光学，Vol．18，No．3，1997，33-36
1.2 Stan Reich, THE USE OF ELECTRO-MECHANICAL MIRROR SCANNING DEVICE, SPIE,Vol. 84, 1976, 47-56
1.3 D. K. Hamiliton, T. Wilson, C. J. Sheppard. Experiment observations of depth discrimation properties of scanning microscopes. Optical Letter, Vol. 6, No. 12, 1983, 625-626
1.4 I. J. Cox, Scanning Optical Fluorescence Microscopy, J. Microscopy, Vol.133, 1984, 149-154
1.5 B. Bertero, Analytic inversion formula for confocal scanning microscopy, J. Opt. Soc. Am. A, Vol. 4, No. 9, 1987, 1748-1750
1.6 T. Wilson, A. R. Carline, Three-dimensional imaging in confocal imaging, J. Microscopy, Vol.149, 1987, 61-66
1.7 K. Carlson, Three-dimensional Microscopy using a Confocal Laser Scanning Microscope, Optical Letters, Vol. 10, No. 2, 1985, 53-55
1.8 Knell Carlson, Niles Aslund, Confocal imaging for 3-d digital Microscopy, Applied Optics, Vol. 26, No. 16, 1987, 3232-3238
1.9 David N. Sitter, William T. Rhode, Three-dimension imaging: a space invariant model for space variant system, Applied Optics, Vol. 29, No. 26, 1990, 3789-3794
1.10 张平，陈德，吴震，黄俊，共焦激光扫描荧光显微镜的扫描系统研究，应用光学，Vol．16，No．3，1995，33-38
1.11 刘峰，刘明，陶纯堪，聂守平，李爱明，激光共焦快速扫描显微成像的实现，应用光学，Vol．16，No．5，1995，27-29
1.12 任雅萍，杨玉孝，赵明涛，王林祥，层析式激光三维测量技术，激光与红外，Vol．27，No．4，1997，226-227
1.13 杨利松，王桂英，徐至展，光学层析成像机理分析，激光与红外，Vol．27，No．4，1997，239-241
1.14 富光华，刘新，实时共焦扫描光学显微镜，半导体光电，Vol．13，No．1，1992，
30-35
1． 15 L. Giniunas, R. Juskaitis, SCANNING FIBER-OPTIC MICROSCOPE, Electronics Letters, Vol.27, No. 9, 1991,724-726
1． 16 M. E. Marhic, S. E. Schscham, M. Epstein, Misalignment in imaging multifibers, Applied Optics, Vol.17, No. 21, 1978, 3503-3507
1． 17 T. Wilson. Images of Phase Edges in Conventional and Scanning Optical Microscopes. Applied Optics, Vol.20, No. 18, 1981, 3238-3244
1． 18 Min Gu, X. Gan, C. J. R. Sheppard, Three-dimensional coherent functions in a fiber-optical confocal scanning microscope, J. Opt. Soc. Am. A, Vol.8, No. 7, 1991, 1019-1025
1． 19 Kenneth P. Ghiggion, Martin R.Harris, Paul G. Spizzirri, Fluorescence lifetime measurements using a novel fiber-optic laser scanning confocal microscope, American Institute of Physics, Vol.66, No.5, 1992, 2999-3002
1． 20 N. Cerre, F. de Fornel, J. P. Goudonnet, Reflection scanning microscope: influence of the distance and the orientation between the fiber and the sample, Optical Society of America, Vol. 10, No.9, 1992, 1909-1917
1． 21 R. Juskaitis, N. Rea, Tony Wilson, Fiber-optic based confocal microscopes using laser detection, Optics Communications, Vol.99, No. 1, 2, 1993, 105-113
1． 22 Arthur F. Gmitro, David Aziz, Confocal microscopy through a fiber-optic imaging bundle, Optics Letters, Vol.18, No. 8, 1993, 565-567
1． 23 Peter M. Delaney, Martin R. Harris, Roger G. King, Fiber-optic laser scanning confocal microscope suitable for fluorescence imaging, Applied Optics, Vol.33, No. 4, 1994, 573-577
1． 24 R. Juskaitis, T. Wilson, Direct-view fiber-optic confocal microscope, Optical Society of America, Vol. 19, No. 22, 1994, 1500-1501
1． 25 David Aziz, Arthur F. Gmitro, Imaging performance of the fiber-optic image-bundle confocal microscope, SPIE, Vol. 2083, 1994
1． 26 N. P. Rea, R. Juskaitis, T. Wilson, Semiconductor laser confocal and interference microscope, Optics Communication, Vol. 125, 1996, 158-167
1.27 宋葵，李增发，光学显微镜的最新发展，物理，Vol．22，No．2，1992，81-86
1.28 魏高尚，向际鹰，吴震，张平，单模光纤共焦扫描显微系统研究，应用光学，Vol．18，No．3，1997，33-36