Three-dimensional image reconstructions from a partially confocal scanning microscope

Jose-Angel Conchello; Qinrong Yu; Jeff W. Lichtman M.D.

doi:10.1117/12.188057

30 September 1994 Three-dimensional image reconstructions from a partially confocal scanning microscope

Jose-Angel Conchello, Qinrong Yu, Jeff W. Lichtman M.D.

Proceedings Volume 2302, Image Reconstruction and Restoration; (1994) https://doi.org/10.1117/12.188057
Event: SPIE's 1994 International Symposium on Optics, Imaging, and Instrumentation, 1994, San Diego, CA, United States

Abstract

Three-dimensional (3-D) microscopy with a non-confocal microscope is fundamentally limited because the optical transfer function (OTF) is zero over a cone-shaped region of the spatial frequency domain and thus the image has a missing cone of optical information that potentially results in artifacts. The strictly confocal scanning microscope (i.e. one with only one pinhole-aperture that is infinitesimally small) does not suffer from this missing cone and thus images are not artifactual. However the pinhole aperture has very low light collection efficiency and images have low signal-to-noise ratio (SNR). Because of this low light-collection efficiency the scanning microscope is commonly operated in a partially confocal regime either by using a larger confocal aperture, multiple confocal apertures working in parallel, or a combination of both. With a larger aperture more out-of-focus fluorescence is collected, with multiple apertures more out-of-focus excitation is produced. In either case the optical axis resolution is degraded relative to that of the ideal confocal microscope. Fortunately neither approach suffers form a missing cone. Frequency components are attenuated relative to the strictly confocal case, but they are no completely missing. We present results that show that applying image restoration methods to partially confocal images it is possible to obtain artifact free images with the same or better resolution than with a strictly confocal microscope.

Citation Download Citation

Jose-Angel Conchello, Qinrong Yu, and Jeff W. Lichtman M.D. "Three-dimensional image reconstructions from a partially confocal scanning microscope", Proc. SPIE 2302, Image Reconstruction and Restoration, (30 September 1994); https://doi.org/10.1117/12.188057

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