FIGURE 3.

Ex vivo Tg mouse wholemount and 6E10 immunofluorescence. Aβ load in the ex vivo retina wholemount was identified by 6E10 immunofluorescence and z-stack confocal imaging taken through the NFL, GCL and IPL. (A) Representative high power confocal image of the NFL demonstrating red immunofluorescence. Note that Aβ specks can be seen in the NFL (white arrowheads), while the fluorescence associated with the RGC cell bodies in the GCL is also seen (yellow arrowheads). (B) As the confocal imaging descends into the wholemount, the GCL is identified in an optical slice by the presence of nuclei, here shown as black circular profiles as the DAPI channel is not shown. Note the fluorescence of the RGCs (yellow arrowheads) is evident in the cytoplasm (shown in boxed inset), and align with the RGCs seen in A. The extracellular Aβ specks can be seen (white arrowheads). (C) An optical slice through the IPL reveals fewer nuclei, as evidence by the lack of black circular nuclear profiles as seen in B. A few Aβ specks (white arrowheads) are seen, but generally less Aβ immunoreactivity is observed in IPL. Scale bar for (A–C) is shown in C = 40 microns. (D) The orthogonal view of the confocal z-stack reconstruction from the Tg retina shown in (A–C). Note red immunoreactivity in the NFL, GCL and IPL (white arrowheads). The immunoreactivity present in the IPL (white arrows) indicates that the antibody penetrated deep into the wholemount tissue. Additional optical slices from this wholemount are shown in Supplementary Figure S2.