Extended Data Fig. 1. Pre-plaque vascular Aβ deposition in App^NL-F mice.

(a, b) Quantification of SPP1 immunoreactivity in CA1 hippocampus (a) versus cerebellum as control region (b), as measured by confocal imaging. 1 datapoint represents 1 ROI per mouse hippocampus, with total of 8 (CA1, a) and 7 (CRB, b) ROI from n = 3 mice per genotype examined over 2 independent experiments. P Values from Two-tailed Mann-Whitney test. Data are shown as Mean ± SEM. (c) Validation of SPP1 antibody (left) and anti-Spp1 probe for smFISH (right) in 3 mo Spp1^KO/KO hippocampal tissue. Scale bar represents 10 µm. Data representative of n = 4 Spp1^KO/KO mice examined over 1 independent experiment. (d) Representative confocal image of Spp1 mRNA expression along GLUT1⁺ vasculature in SLM of 6 mo App^NL-F mice as characterized by smFISH-IHC. Scale bar represents 20 µm. Insert: 3D reconstruction. Scale bar represents 5 µm. Data are representative of 3 App^NL-F mice examined over at least 4 independent experiments. (e) Representative confocal images showing 6E10 plaque staining in hippocampus of 6 mo versus 15 mo App^NL-F mice. Scale bar represents 100 µm. Data representative of n = 3 App^NL-F mice mice per time point, examined over 2 independent experiments. (f) Representative confocal images showing oAβ around GLUT1⁺ blood vessels as characterized by NAB61 immunoreactivity in 6 mo WT versus 6 mo App^NL-F SLM. Scale bar represents 10 µm. Data representative of n = 3 mice per genotype, examined over 2 independent experiments. (g) Representative confocal images showing vascular oAβ deposition using alternative antibodies 4G8 and HJ3.4 that recognize Aβ17-24 and Aβ1-13, respectively, in 6 mo App^NL-F SLM. Scale bar represents 10 µm. Data representative of n = 3 App^NL-F mice examined over 2 independent experiments. (h) Representative confocal images showing vascular oAβ (NAB61) in 15 mo WT versus App^NL-F SLM. Scale bar represents 20 µm.

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