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Quantification of collagen fiber structure using second harmonic generation imaging and two-dimensional discrete Fourier transform analysis: Application to the human optic nerve head

DOI: 10.1002/jbio.201800376 DOI Help

Authors: Jacek K. Pijanka (Cardiff University) , Petar P. Markov (Cardiff University) , Dan Midgett (Johns Hopkins University) , Neil G. Paterson (Diamond Light Source) , Nick White (Cardiff University) , Emma J. Blain (Cardiff University) , Thao Nguyen (Johns Hopkins University) , Harry A. Quigley (Johns Hopkins University) , Craig Boote (University of Cardiff)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biophotonics , VOL 78

State: Published (Approved)
Published: January 2019
Diamond Proposal Number(s): 14757

Abstract: Second harmonic generation (SHG) microscopy is widely used to image collagen fiber microarchitecture due to its high spatial resolution, optical sectioning capabilities and relatively nondestructive sample preparation. Quantification of SHG images requires sensitive methods to capture fiber alignment. This article presents a two‐dimensional discrete Fourier transform (DFT)–based method for collagen fiber structure analysis from SHG images. The method includes integrated periodicity plus smooth image decomposition for correction of DFT edge discontinuity artefact, avoiding the loss of peripheral image data encountered with more commonly used windowing methods. Outputted parameters are as follows: the collagen fiber orientation distribution, aligned collagen content and the degree of collagen fiber dispersion along the principal orientation. We demonstrate its application to determine collagen microstructure in the human optic nerve head, showing its capability to accurately capture characteristic structural features including radial fiber alignment in the innermost layers of the bounding sclera and a circumferential collagen ring in the mid‐stromal tissue. Higher spatial resolution rendering of individual lamina cribrosa beams within the nerve head is also demonstrated. Validation of the method is provided in the form of correlative results from wide‐angle X‐ray scattering and application of the presented method to other fibrous tissues.

Journal Keywords: collagen fiber structure; discrete Fourier transform; edge effect artefact correction; nonlinear microscopy; optic nerve head; second harmonic generation

Subject Areas: Biology and Bio-materials


Instruments: I03-Macromolecular Crystallography