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Journal of Histochemistry and Cytochemistry, Vol. 50, 1469-1474, November 2002, Copyright © 2002, The Histochemical Society, Inc.

ARTICLE

A Quantitative Method to Determine the Orientation of Collagen Fibers in the Dermis

Maril L. Noorlandera, Paris Melisb, Ard Jonkera, and Cornelis J.F. Van Noordena
a Academic Medical Center, University of Amsterdam, Departments of Cell Biology and Histology, Amsterdam, The Netherlands
b Plastic, Reconstructive and Handsurgery, Amsterdam, The Netherlands

Correspondence to: Cornelis J.F. Van Noorden, Dept. of Cell Biology and Histology, Academic Medical Center, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands. E-mail: c.j.vannoorden@amc.uva.nl

We have developed a quantitative microscopic method to determine changes in the orientation of collagen fibers in the dermis resulting from mechanical stress. The method is based on the use of picrosirius red-stained cryostat sections of piglet skin in which collagen fibers reflect light strongly when epipolarization microscopy is used. Digital images of sections were converted into binary images that were analyzed quantitatively on the basis of the length of the collagen fibers in the plane of the section as a measure for the orientation of the fibers. The length of the fibers was expressed in pixels and the mean length of the 10 longest fibers in the image was taken as the parameter for the orientation of the fibers. To test the procedure in an experimental setting, we used skin after 0 and 30 min of skin stretching. The orientation of the fibers in sections of control skin differed significantly from the orientation of fibers in sections of skin that was stretched mechanically for 30 min [76 ± 15 (n=5) vs 132 ± 36 (n=5)]. The method described here is a relatively simple way to determine (changes in) the orientation of individual collagen fibers in connective tissue and can also be applied for analysis of the orientation of any other structural element in tissues so long as a representative binary image can be created.

(J Histochem Cytochem 50:1469–1474, 2002)

Key Words: image analysis, quantitative microscopy, picrosirius red staining, collagen orientation, dermis, wound closure


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