Figure 5.

In situ binding assay in Tg2576 brain sections. Sixteen month-old Tg2576 mice were perfused with PBS and brains were post-fixed in 4% paraformaldehyde. Coronal brain sections (40 μm thick) were washed three times with PBS and incubated with various concentrations of test compounds (4 sections/concentration; 6 concentrations/compound) at room temperature for 30 min. Brain sections were washed twice with PBS and 50% ethanol in PBS then cover-slipped. A. Representative images of fluorescent labeling with resorufin, ethoxy-resorufin (Ethoxy), benzyloxy-resorufin (Benzyloxy), and methoxy-X34 (X34) are seen. Note that all resorufin derivatives selectively bind CAA-laden arterioles (arrowheads), whereas the methoxy-X34 labels both CAA and neuritic plaques (arrows). Scale bar: 100 μm. Fluorescent images of the sensorimotor cortex were taken and the fluorescent intensity on CAA (B) and neuritic plaques (C) was quantified. Saturation binding curves were plotted to obtain the binding affinity (K_D) values (see Table 2).