Table 1.
DNA sequence for the β55 aptamer including the T7 polymerase primer, highlighted in bold text.
Figure 1.
Predicted Secondary Structure of RNA Aptamers.
Predicted secondary structure of the β55 (left) and β55rc (right) aptamer probes with base pair probability indicated by the color scale bar. Minimum free energy structures were determined using the RNAfold Webserver suite of programs.
Figure 2.
β55 Staining of Amyloid Plaques in Ex Vivo Human AD Brain Tissue.
Merged red and green channel confocal images of frozen-section human AD brain tissue stained with biotinylated β55 (a) and β55rc (b). β55 positive plaques (green) were clearly visible, while only a few very faint β55rc positive plaques were observed. Background auto-fluorescence, observed in both red and green channels, is shown in yellow. (c) Fluorescence images of human AD brain tissue costained with biotinylated-β55 (red) and Thioflavin-S (green). β55 colocalized with Thioflavin-S positive plaques. (Scale bars: 50 µm).
Figure 3.
β55 Staining of Dot Blots of Synthetic Aβ Aggregates.
(a) Dot blot of synthetic Aβ1–42 and Aβ1–40 aggregates probed with biotinylated-β55. (b) Western blot of the synthetic Aβ1–42 and Aβ1–40 aggregates probed with 6E10 antibody. The increased staining of Aβ1–42 aggregates in the dot blot relative to Aβ1–40 aggregates is consistent with the greater fibril and high molecular weight oligomer composition of Aβ1–42 aggregates observed in the western blot.
Figure 4.
In Vivo Imaging of β55 Positive Amyloid Plaques.
In vivo 2-photon microscopy images from an 18 month old APP/PS1 transgenic mouse obtained 1 hour after topical application of fluorescein-labeled β55 (a,b). Texas Red labeled dextran was intravenously injected for visualization of blood vessels. β55 positive plaques and cerebral amyloid angiopathy are clearly visible in the cortex (a) and vasculature (b), respectively. (Scale bars: 20 µm).
Figure 5.
Colocalization of β55 and Methoxy-XO4 Positive Amyloid Plaques.
In vivo 2-photon microscopy plaque images from a 7 month old APP/PS1 transgenic mouse acquired 1 hour after topical application of fluorescein-labeled β55 (a,d) and 1 day after intraperitoneal injection of methoxy-XO4 (b,e). While methoxy-XO4 stains only the dense core of the plaque, β55 stains both the plaque core and a diffuse halo surrounding the plaque (c,f). (Scale bars: 20 µm).
Figure 6.
Contrast-to-Noise Ratio for β55 and β55rc Positive Amyloid Plaques.
(a) Representative in vivo 2-photon microscopy images from 7.5 month old APP/PS1 transgenic mice acquired 1 hour (left column) and 24 hours (right column) after topical application of either fluorescein-labeled β55 (top row) or β55rc (bottom row). Most β55 plaques were still visible 24 hours after topical application. In contrast, only a small number of very faint β55rc plaques were still visible after 24 hours. (b) Average plaque contrast-to-noise ratio (CNR) observed 1 hour and 24 hours following topical application of fluorescein-labeled β55 (n = 2) or β55rc (n = 2). β55 positive plaques had a significantly greater CNR than β55rc plaques (p<0.01) at both time points. (Scale bars: 50 µm).
Table 2.
Contrast-to-noise ratio (CNR) distribution of amyloid plaques observed in 2-photon in vivo images of 7.5 month-old APP/PS1 transgenic mice with β55 and β55rc.