Measurement-based depth of focus

Stephanie L. Ellenberger; Ian T. Young

doi:10.1117/12.273648

2 May 1997 Measurement-based depth of focus

Stephanie L. Ellenberger, Ian T. Young

Proceedings Volume 2982, Optical Diagnostics of Biological Fluids and Advanced Techniques in Analytical Cytology; (1997) https://doi.org/10.1117/12.273648
Event: BiOS '97, Part of Photonics West, 1997, San Jose, CA, United States

Abstract

An important point in the automated evaluation of molecular cytogenetic preparations is focusing the specimen before taking images. The in-focus position can vary for different positions on the slide, thus every new field of view has to be refocused. Another problem is that objects in one field of view can lie in different planes with the effect that one object is out-of-focus while the other one is in-focus. We want to investigate the importance of focus with regard to measurements like object intensity. Diffraction wave theory leads to a formula, which can be used to determine the depth-of-focus (DOF), the tolerance in the in-focus position where the image is still sharp. However practice shows that it is possible to get good measurement results even if the specimen is not imaged perfectly.the relation between the theoretical and practical DOF is analyzed. The objects under investigation are either 2D or 3D and are imaged in absorption and fluorescence mode. It appears that the DOF based on measurements can be 5 to 10 times bigger than a DOF based on wave theory with only little loss of accuracy.

Citation Download Citation

Stephanie L. Ellenberger and Ian T. Young "Measurement-based depth of focus", Proc. SPIE 2982, Optical Diagnostics of Biological Fluids and Advanced Techniques in Analytical Cytology, (2 May 1997); https://doi.org/10.1117/12.273648

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