(A and B) Amyloid plaques were labeled 24 hours before with methoxy-X04 (MeX04). Mice were then anesthetized, and a fluorescent anti-Aβ antibody was injected intracisternally. Mice received either i.p. isotonic saline (Control) or hypertonic saline (+HTS) at the onset of the intracisternal infusion. After 120 minutes, mice were perfusion fixed with a fluorescent lectin to label the vasculature. (C) Representative ex vivo images of intact brains upon removal from the cranium (bottom left; scale bar: 2 mm) and after coronal sectioning to evaluate antibody penetrance into the brain (top; scale bar: 500 μm). Confocal images of the antibody and Aβ plaques (arrowheads) surrounding the perivascular spaces of penetrating arteries (bottom right; scale bar: 100 μm). (D) Quantification of ex vivo coronal section Aβ antibody fluorescence MPI (mean ± SEM; n = 5 mice/group; unpaired 2-tailed t test; **P = 0.0039). (E) Representative high-magnification confocal images of perivascular Aβ plaques (scale bar: 20μm). (F) Percentage of target engagement shown by colabeling of the antibody with MeX04⁺ Aβ plaques (mean ± SEM; n = 5 mice/group; unpaired t test; **P = 0.005). (G) Nearest neighbor analysis of the average distance of a colabeled plaque from its nearest perivascular space (PVS) in μm (mean ± SEM; total number of colabeled plaques/number of mice in group; unpaired t test; ****P < 0.0001). (H) Histogram and cumulative frequency plot of the number of colabeled plaques and distance from the nearest PVS. (I) Representative high-magnification confocal image with orthogonal views showing the anti-Aβ antibody engaging the surface of a plaque (arrows). Scale bar: 20 μm. (J) Three-dimensional reconstruction of Aβ plaques from an +HTS-treated mouse showing antibody targeting and engaging plaque surface (scale bar: 20 μm [both images]). (K) Plaque burden was the same between groups (mean ± SEM; n = 5 mice/group; unpaired t test; P = 0.6165).