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A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration

Colour deconvolution is a method used in diagnostic brightfield microscopy to transform colour images of multiple stained biological samples into images representing the stain concentrations. It is applied by decomposing the absorbance values of stain mixtures into absorbance values of single stains...

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Autores principales: Haub, Peter, Meckel, Tobias
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4519787/
https://www.ncbi.nlm.nih.gov/pubmed/26223691
http://dx.doi.org/10.1038/srep12096
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author Haub, Peter
Meckel, Tobias
author_facet Haub, Peter
Meckel, Tobias
author_sort Haub, Peter
collection PubMed
description Colour deconvolution is a method used in diagnostic brightfield microscopy to transform colour images of multiple stained biological samples into images representing the stain concentrations. It is applied by decomposing the absorbance values of stain mixtures into absorbance values of single stains. The method assumes a linear relation between stain concentration and absorbance, which is only valid under monochromatic conditions. Diagnostic applications, in turn, are often performed under polychromatic conditions, for which an accurate deconvolution result cannot be achieved. To show this, we establish a mathematical model to calculate non-monochromatic absorbance values based on imaging equipment typically used in histology and use this simulated data as the ground truth to evaluate the accuracy of colour deconvolution. We show the non-linear characteristics of the absorbance formation and demonstrate how it leads to significant deconvolution errors. In particular, our calculations reveal that polychromatic illumination causes 10-times higher deconvolution errors than sequential monochromatic LED illumination. In conclusion, our model can be used for a quantitative assessment of system components - and also to assess and compare colour deconvolution methods.
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spelling pubmed-45197872015-08-05 A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration Haub, Peter Meckel, Tobias Sci Rep Article Colour deconvolution is a method used in diagnostic brightfield microscopy to transform colour images of multiple stained biological samples into images representing the stain concentrations. It is applied by decomposing the absorbance values of stain mixtures into absorbance values of single stains. The method assumes a linear relation between stain concentration and absorbance, which is only valid under monochromatic conditions. Diagnostic applications, in turn, are often performed under polychromatic conditions, for which an accurate deconvolution result cannot be achieved. To show this, we establish a mathematical model to calculate non-monochromatic absorbance values based on imaging equipment typically used in histology and use this simulated data as the ground truth to evaluate the accuracy of colour deconvolution. We show the non-linear characteristics of the absorbance formation and demonstrate how it leads to significant deconvolution errors. In particular, our calculations reveal that polychromatic illumination causes 10-times higher deconvolution errors than sequential monochromatic LED illumination. In conclusion, our model can be used for a quantitative assessment of system components - and also to assess and compare colour deconvolution methods. Nature Publishing Group 2015-07-30 /pmc/articles/PMC4519787/ /pubmed/26223691 http://dx.doi.org/10.1038/srep12096 Text en Copyright © 2015, Macmillan Publishers Limited http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Haub, Peter
Meckel, Tobias
A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration
title A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration
title_full A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration
title_fullStr A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration
title_full_unstemmed A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration
title_short A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration
title_sort model based survey of colour deconvolution in diagnostic brightfield microscopy: error estimation and spectral consideration
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4519787/
https://www.ncbi.nlm.nih.gov/pubmed/26223691
http://dx.doi.org/10.1038/srep12096
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