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. Author manuscript; available in PMC: 2021 Apr 7.
Published in final edited form as: J Comp Neurol. 2018 Dec 4;527(13):2122–2145. doi: 10.1002/cne.24555

Figure 4. Methoxy-X04 accurately labels the location of plaques but underestimates plaque density.

Figure 4.

(a-c) Automated segmentation results (a), IHC for Aβ (6E10 antibody) (b), and an overlay of the automated segmentation and thresholded IHC image from the same section (c) are shown for one section from each mouse line. APP/PS1 and hAPP-J20 brains are from 13-month-old mice. The Tg2576 brain is from a 19-month-old mouse. In the segmentation images in (a), red indicates segmented plaques, and gray the putative plaque signal that was removed by the adaptive filtering step in the segmentation algorithm. Automatically detected tissue edges are also colored gray. Image alignment in the overlay images (c) is imperfect due to tissue deformation during antibody labeling. (d) maximum intensity projection of a confocal image stack through the center of one plaque for each mouse line. Upper grayscale image shows 6E10 labeling; bottom color image shows 6E10 labeling overlaid with methoxy-X04 and DAPI fluorescence (both in cyan). Arrowheads indicate the methoxy-X04 positive core of each plaque. Scale = 500 μm (a-c), 50 μm (d). (e) Aβ antibody-labeled plaque density and segmented methoxy-X04-labeled plaque density plotted for each ROI measured (log scale). Points are colored by the Allen CCFv3 reference atlas region in which each ROI was drawn (isocortex, hippocampal formation, cortical subplate, olfactory areas, striatum, and thalamus). Inset shows a coronal section from the Allen CCFv3 Reference Atlas with region colors. The shape of each point indicates which mouse line it belongs to: circles for APP/PS1, squares for Tg2576, and triangles for hAPP-J20. Methoxy-X04 density was lower than Aβ antibody-labeled density for every point, but the two measurements were highly correlated (Pearson’s r = 0.95, p<0.0001). (f) The fraction of Aβ antibody-labeled plaque density that was detected by automated segmentation was significantly lower for hAPP-J20 mice compared to APP/PS1 mice. (g) For individual plaques, the ratio of methoxy-X04 labeling to Aβ antibody labeling is plotted for each mouse line. hAPP-J20 mice had significantly lower fractions of methoxy-X04/Aβ antibody labeling.