Stereo pair showing sheets.

<p>Sheets located just beneath the subserosal sheet are indicated by the green bar, while the deeper layer of smaller fasciculi is indicated by the black bar at the top of the image. The marked difference in structure between the sheets under the green bar and fasciculi under the black bar points to a clear distinction between the two layers. Structures were highlighted at random locations to highlight the large structures shown here (for details, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0173404#pone.0173404.s019" target="_blank">S1 Appendix</a>). Right-hand panel shows the global position of the images. Scale bars represent 2 mm.</p

Empirical CDFs of the distribution of bundle widths.

<p>Widths in the human tissue block are markedly higher than those in the rat tissue blocks, as indicated by the shift to the right of the distribution. Additionally, the third rat tissue block appears to have smaller widths than the other two rat tissue blocks, as indicated by a shift to the left of the distribution.</p

Comparison with anisotropy maps.

<p>DT MRI was simulated on the reconstruction to obtain fractional anisotropy (<b>A</b>) and diffusion tensor ellipses (<b>B</b>) at a resolution of 380 <i>μ</i>m per voxel (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0173404#pone.0173404.s019" target="_blank">S1 Appendix</a> for detailed methods). These can be directly compared to the original weighting (<b>C</b>) and directions (<b>D</b>), which are at a resolution of 47.5 <i>μ</i>m per voxel. The simulated diffusion tensor images are similar to those presented by Weiss <i>et al.</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0173404#pone.0173404.ref003" target="_blank">3</a>].</p

Extracting nuclei from histological slides.

<p><b>A</b>: A 500 × 500 pixel region of tissue. <b>B</b> & <b>C</b>: Local red and blue thresholds respectively, found by considering the average values within 32 × 32 pixel regions. <b>D</b>: Binary image obtained by applying these thresholds to the original image <b>A</b>. <b>E</b>: Ellipses found by applying the ImageJ “Analyze Particles” function [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0173404#pone.0173404.ref020" target="_blank">20</a>] to <b>D</b>, superimposed onto a lower intensity copy of <b>A</b>, showing the nuclei isolated from the rest of the tissue. Scale bars represent 50 <i>μ</i>m.</p

Scalar representation of two-dimensional vectors.

<p><b>A</b>: A representation of the directions vectors, where red and green indicate the direction of planar vectors, and blue indicates vertical pixels. <b>B</b>: The grey image generated from smoothing the nuclear counts present in the vertical pixels. <b>C</b>: The grey image generated from the planar vectors represented in <b>A</b>. Scale bars represent 1 mm.</p

The varying degrees of heterogeneity within a slide.

<p>The lines represent directions of <i>planar</i> pixels. Each image shows the planar directions in a 16 × 16 pixel (approx. 1 × 1 mm²) tile. <b>A</b>: An example of a tile of high heterogeneity. The large amount of empty space and variation in fascicular direction allows for a precise registration of the tile. <b>B</b>: An example of a tile of low heterogeneity. Here the tile comprises vectors with little variation in direction. This homogeneity can readily lead to an unreliable registration, and accordingly, the area of interest needs to be expanded to obtain a higher level of heterogeneity.</p

Median bundle width for each tissue block.

<p>Median bundle width for each tissue block.</p

Number of slides at each step of processing.

<p>Number of slides at each step of processing.</p

Variability of log-transformed bundle widths in registered and unregistered slides.

<p><b>R1</b>, <b>R2</b>, and <b>R3</b> represent each of the rat tissue blocks, <b>H</b> represents the human tissue block. The general similarity in shape between registered and unregistered slides in each block suggests that any deformation incurred by the registration process has little effect on the overall structure of the tissue block.</p

Difference in mean log-transformed bundle widths between registered and unregistered tissue blocks by slide.

<p>Dashed lines indicate a 5% change in width. The vast majority of points lie within the 5% bounds for each block, suggesting that even at an individual slide level the deformation is not significant.</p