Shading Correction: Compensation for Illumination and Sensor Inhomogeneities

Ian T. Young1

1 Delft University of Technology, Delft, The Netherlands
Publication Name:  Current Protocols in Cytometry
Unit Number:  Unit 2.11
DOI:  10.1002/0471142956.cy0211s14
Online Posting Date:  May, 2001
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Abstract

The interaction between objects in real space, the illumination, and the camera frequently leads to a situation in which a microscope image exhibits significant shading across the field of view. In general this shading effect is undesirable and requires elimination, especially for quantitative microscopy. Starting with a simple mathematical model, this unit develops procedures to correct images for the shading thus introduced in the imageā€formation process. Two cases are distinguished: the a priori, which assumes the availability of calibration images in addition to the images of interest; and the a posteriori, which assumes that only the recorded images of interest are available.

     
 
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Table of Contents

  • Flat and Non‐Flat Fields
  • Shading Correction
  • Results
  • Conclusions
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

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Figures

Videos

Literature Cited

Literature Cited
   Castleman, K.R. 1996. Digital Image Processing (2nd ed.). Prentice‐Hall, Englewood Cliffs, N.J.
  Dr. Young wishes to acknowledge the support of the Rolling Grants program of the Foundation for Fundamental Research in Matter (FOM) and the Delft Inter‐Faculty Research Center (DIOC) program “Intelligent Molecular Diagnostic Systems.”
   Frisby, J.P. 1980. Seeing: Illusion, Brain and Mind. Oxford University Press, Oxford.
   Giardina, C.R. and Dougherty, E.R. 1988. Morphological Methods in Image and Signal Processing. Prentice‐Hall, Englewood Cliffs, N.J.
   Heijmans, H.J.A.M. 1994. Morphological Image Operators. Academic Press, Boston.
   Murch, G.M. 1973. Visual and Auditory Perception. Bobbs‐Merrill, New York.
   Oppenheim, A.V., Schafer, R.W., and Stockham, T.G. Jr. 1968. Non‐linear filtering of multiplied and convolved signals. Proc. IEEE 56:1264‐1291.
   Stockham, T.G. 1972. Image processing in the context of a visual model. Proc. IEEE 60:828‐842.
   v.d. Doel, L.R., Klein, A.D., Ellenberger, S.L., Netten, H., Boddeke, F.R., van Vliet, L.J., and Young, I.T. 1998. Quantitative evaluation of light microscopes based on image processing techniques. Bioimaging 6:138‐149.
   Young, I.T. 1989. Image fidelity: Characterizing the imaging transfer function. In Fluorescence Microscopy of Living Cells in Culture: Quantitative Fluorescence Microscopy–Imaging and Spectroscopy (D.L. Taylor and Y.L. Wang, eds.) pp. 1‐45. Academic Press, San Diego.
   Young, I.T., Gerbrands, J.J., and van Vliet, L.J. 1998. Image processing fundamentals. In The Digital Signal Processing Handbook (V.K. Madisetti and D.B. Williams, eds.) pp. 51.1‐51.81. CRC Press in cooperation with IEEE Press, Boca Raton, Fla.
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