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Analytical Cellular Pathology
Volume 20, Issue 1, Pages 7-15
http://dx.doi.org/10.1155/2000/459351

Reconstruction of Axial Tomographic High Resolution Data from Confocal Fluorescence Microscopy: A Method for Improving 3D FISH Images

R. Heintzmann,1 G. Kreth,1,2 and C. Cremer1,2

1Applied Optics and Information Processing, Institute of Applied Physics, University of Heidelberg, Albert‐Ueberle Str. 3‐5, D‐69120 Heidelberg, Germany
2Interdisciplinary Center for Scientific Computing (IWR), University of Heidelberg, Germany

Accepted 31 March 2000

Copyright © 2000 Hindawi Publishing Corporation. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Fluorescent confocal laser scanning microscopy allows an improved imaging of microscopic objects in three dimensions. However, the resolution along the axial direction is three times worse than the resolution in lateral directions. A method to overcome this axial limitation is tilting the object under the microscope, in a way that the direction of the optical axis points into different directions relative to the sample. A new technique for a simultaneous reconstruction from a number of such axial tomographic confocal data sets was developed and used for high resolution reconstruction of 3D‐data both from experimental and virtual microscopic data sets. The reconstructed images have a highly improved 3D resolution, which is comparable to the lateral resolution of a single deconvolved data set. Axial tomographic imaging in combination with simultaneous data reconstruction also opens the possibility for a more precise quantification of 3D data. The color images of this publication can be accessed from http://www.esacp.org/acp/2000/20‐1/heintzmann.htm. At this web address an interactive 3D viewer is additionally provided for browsing the 3D data. This java applet displays three orthogonal slices of the data set which are dynamically updated by user mouse clicks or keystrokes.