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

Haub, Peter and Meckel, Tobias (2015):
A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration.
In: Scientific reports, p. 12096, 5, ISSN 2045-2322, [Article]

Abstract

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.

Item Type: Article
Erschienen: 2015
Creators: Haub, Peter and Meckel, Tobias
Title: A Model based Survey of Colour Deconvolution in Diagnostic Brightfield Microscopy: Error Estimation and Spectral Consideration.
Language: English
Abstract:

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.

Journal or Publication Title: Scientific reports
Volume: 5
Divisions: 10 Department of Biology
10 Department of Biology > Membrane Dynamics
Date Deposited: 04 Aug 2015 06:06
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