The impact of the depth of field on cytogenetic image quality in scanning microscopy
2011
The purpose of this study is to investigate the impact of the depth of field (DOF) of microscopic systems on cytogenetic
image qualities. Due to the narrow DOF of high magnification, large numerical aperture (N.A.) objective lenses, random
vibrations of even high precision scanning stages may result in large amount of off focused images. In this study, the
DOF of microscopic systems with various objective magnifications/numerical apertures (N.A.) is first measured using
standard resolution targets. The impact of DOF on cytogenetic image qualities is then subjectively evaluated with
clinical samples, by comparing the band shape and sharpness of analyzable chromosomes. For a specific digital
microscopic system with 100× objective lens (N.A. = 1.25), the results of observational studies revealed that
chromosomal bands are still recognizable when the images are obtained approximately +/- 1 μm from the focusing plane.
The chromosomal bands become fuzzy and unrecognizable when the system is 1.5 μm away from the focusing position.
The results of this preliminary experimental study may provide useful design trade-off parameters for developing
optimal scanning microscopic systems for cytogenetic applications.
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