Confocal Laser Scanning Microscopy

Abstract

, 75 pages Optical imaging techniques have improved much over the last fifty years since the invention of the laser. With a high brightness source many imaging applications which were once inaccessible to researchers have now become a reality. Among these techniques, the most beneficial one is the use of lasers for both wide-field and confocal imaging systems. The aim of this study was to design a laser imaging system based on the concept of laser scanning confocal microscopy. Specifically the optical system was based on optical fibers allowing the user to image remote areas such as the inner surface of rifled gun barrels and/or pipes with a high degree of precision (+/-0.01 mm). In order to build such a system, initially the theoretical foundation for a confocal as well as a wide-field imaging system was analyzed. Using this basis a free-space optical confocal system was built and analyzed. The measurements support the fact that both the objective numerical aperture and pinhole size play an important role in the radial and axial resolution of the system as well as the quality of the images obtained. v To begin construction of a confocal, optical-fiber based imaging system first an allfiber wide-field imaging system was designed and tested at a working wavelength of 1550 nm. Then an all fiber confocal system was designed at a working wavelength of 808 nm. In both cases results showed that while lateral resolution was adequate, axial resolution suffered since it was found that the design of the optical system needs to take into account under-filling of the objective lens, a result common with the use of laser beams whose divergence is not at all like that of a point source. The work done here will aid technology that will be used in the elimination process of faulty rifling fabrication in defense industry. The reason why the confocal technique is preferred to the conventional wide-field one is the need for better resolution in all directions. Theoretical concepts and mathematical background are discussed as well as the experimental results and the practical advantages of such a system. Keywords: Confocal-scanning laser microscope, wide-field imaging techniques, fiber optical imaging techniques Halil Berberoğlu, for his contributions especially to the MATLAB work. The author would also like to thank to her mother, father and little brother for their invaluable devotion and encouragement, and her fiancé Ulaş for his great understanding and patience. The fellowships of my colleagues and friends are gratefully acknowledged.

An LED-Based structured illumination microscope using a digital micromirror device and GPU accelerated image reconstruction

“…In order to achieve resolution improvements beyond the diffraction limit, a number of super-resolution imaging techniques have been developed, i.e. photoactivated localization microscopy (PALM), stochastic optical reconstruction microscopy (STORM), stimulated emission depletion microscopy (STED) and structured illumination microscopy (SIM) [3][4][5][6][7]. In addition to these techniques, the anisotropic resolution of the 2D camera detector can be used to improve the spatial resolution of microscopic images [8].…”

A review of advanced microscopy techniques for the development of nanotechnology in agriculture, food, and the environment

“…As an improvement in conventional microscopic techniques, confocal laser scanning microscopy (CLSM) has emerged as a technique for resolving the structure of many samples through the internalization of fluorescent molecules (Olivier & Moine, 2013). CLSM microscopy (figure 4a) is a technique that conjugates focal planes of specimens between objective lens and detectors to sample visualization.…”