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Originally published as Biophys J. BioFAST on May 13, 2005.
doi:10.1529/biophysj.104.053652
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Biophysical Journal 89:1302-1307 (2005)
© 2005 The Biophysical Society

Determining Diffusion Coefficients in Inhomogeneous Tissues Using Fluorescence Recovery after Photobleaching

Y. H. Sniekers and C. C. van Donkelaar

Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands

Correspondence: Address reprint requests to René C. C. van Donkelaar, Eindhoven University of Technology, Biomedical Engineering, PO Box 513, 5600 MB Eindhoven, The Netherlands; Tel.: NL-040-2473135; E-mail: c.c.v.donkelaar{at}tue.nl.

Diffusion plays an important role in the transport of nutrients and signaling molecules in cartilaginous tissues. Diffusion coefficients can be measured by fluorescence recovery after photobleaching (FRAP). Available methods to analyze FRAP data, however, assume homogeneity in the environment of the bleached area and neglect geometrical restrictions to diffusion. Hence, diffusion coefficients in inhomogeneous materials, such as most biological tissues, cannot be assessed accurately. In this study, a new method for analyzing data from FRAP measurements has been developed, which is applicable to inhomogeneous tissues. It is based on a fitting procedure of the intensity recovery after photobleaching with a two-dimensional finite element analysis, which includes Fick's law for diffusion. The finite element analysis can account for distinctive diffusivity in predefined zones, which allows determining diffusion coefficients in inhomogeneous samples. The method is validated theoretically and experimentally in both homogeneous and inhomogeneous tissues and subsequently applied to the proliferation zone of the growth plate. Finally, the importance of accounting for inhomogeneities, for appropriate assessment of diffusivity in inhomogeneous tissues, is illustrated.




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