From Image to Data Using Common Image‐Processing Techniques

Laura R. Sysko1, Michael A. Davis1

1 Nikon Instruments Inc., Melville, New York
Publication Name:  Current Protocols in Cytometry
Unit Number:  Unit 12.21
DOI:  10.1002/0471142956.cy1221s54
Online Posting Date:  October, 2010
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Abstract

A digital microscopy image is an array of number values, which with adequate contrast can be interpreted as spatial information. Through processing and analysis by mathematical means, using computer‐assisted imaging software programs, raw image data contrast can be enhanced to improve the extraction of image features for measurement and analysis. This mathematical feature extraction (referred to as segmentation) provides the basis for general image processing. The methods discussed in this unit address common image analysis challenges such as object counting with touching objects, objects within other objects, and object identification in a field with uneven illumination or uneven brightness, along with step‐by‐step procedures for achieving these results. Curr. Protoc. Cytom. 54:12.21.1‐12.21.17. © 2010 by John Wiley & Sons, Inc.

Keywords: image analysis; segmentation; image processing; image measurement; digital imaging; thresholding; automated counting

     
 
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Table of Contents

  • Introduction
  • Image Anatomy
  • Image Processing
  • Concluding Remarks
  • Literature Cited
  • Figures
     
 
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Materials

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Figures

Videos

Literature Cited

Literature Cited
   Cox, P.W., Paul, G.C., and Thomas, C.R. 1998. Image analysis of the morphology of filamentous micro‐organisms. Microbiology 144:817‐827.
   Hinchcliffe, E.H. 2003. The use and manipulation of digital image files in light microscopy. In Methods in Cell Biology, Vol. 72. Digital Microscopy: A 2nd ed. of Video Microscopy (G. Sluder and D.E. Wolf, eds.) pp. 271‐288. Elsevier Academic Press, San Diego.
   Inoué, T. and Gliksman, N. 2003. Optimizing microscopy and analysis. In Methods in Cell Biology, Vol. 72. Digital Microscopy: A 2nd ed. of Video Microscopy (G. Sluder and D.E. Wolf, eds.) pp. 243‐270. Elsevier Academic Press, San Diego.
   Inoué, S. and Spring, K.R. 1997. Digital image processing. In Video Microscopy: The Fundamentals, 2nd ed. pp. 509‐558. Plenum Press, New York.
   Pawley, J.B. 2006a. Points, pixels, and gray levels: Digitizing image data. In Handbook of Biological Confocal Microscopy (J.B. Pawley, ed.) pp. 59‐79. Springer‐Science+Business Media, New York.
   Pawley, J.B. 2006b. More than you ever really wanted to know about charged‐coupled devices. In Handbook of Biological Confocal Microscopy (J.B. Pawley, ed.) pp. 918‐931. Springer‐Science+Business Media, New York.
   Russ, J.C. 1995. The Image Processing Handbook, 2nd ed. CRC Press, Boca Raton, Fla.
   Waters, J.C. 2009. Accuracy and precision in quantitative fluorescence microscopy. J. Cell Biol. 185:1135‐1148.
   Zwier, J.M., Van Rooij, G.J., Hofstraat, J.W., and Brajenhoff, G.J. 2004. Image calibration in fluorescence microscopy. J. Microsc. 216:15‐24.
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