5 May 1994 Intercellular fluorescence background on microscope slides: some problems and solutions for automatic analysis
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Although high contrast between signal and the dark background is often claimed as a major advantage of fluorescence staining in cytology and cytogenetics, in practice this is not always the case and in some circumstances the inter-cellular or, in the case of metaphase preparations, the inter-chromosome background can be both brightly fluorescent and vary substantially across the slide or even across a single metaphase. Bright background results in low image contrast, making automatic detection of metaphase cells more difficult. The background correction strategy employed in automatic search must both cope with variable background and be computationally efficient. The method employed in a fluorescence metaphase finder is presented, and the compromises involved are discussed. A different set of problems arise when the analysis is aimed at accurate quantification of the fluorescence signal. Some insight into the nature of the background in the case of comparative genomic hybridization is obtained by image analysis of data obtained from experiments using cell lines with known abnormal copy numbers of particular chromosome types.
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Jim Piper, Jim Piper, Damir Sudar, Damir Sudar, Don Peters, Don Peters, Daniel Pinkel, Daniel Pinkel, "Intercellular fluorescence background on microscope slides: some problems and solutions for automatic analysis", Proc. SPIE 2173, Image Acquisition and Scientific Imaging Systems, (5 May 1994); doi: 10.1117/12.175170; https://doi.org/10.1117/12.175170

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