Journal: Molecular Therapy. Methods & Clinical Development

Article Title: In-depth comparison of Anc80L65 and AAV9 retinal targeting and characterization of cross-reactivity to multiple AAV serotypes in humans

doi: 10.1016/j.omtm.2023.05.016

Figure Lengend Snippet: Anc80 vector has increased tropism for photoreceptors and RPE compared with AAV9 after subretinal injection in WT C57Bl/6J mice Representative immunofluorescent images of retinas comparing AAV9.CB.GFP (n = 4), Anc80.CB.GFP (n = 3), AAV9.546.GFP (n = 3), and Anc80.546.GFP (n = 3) transduction after subretinal injection. (A) Large-Scan 30X stitched image of whole retina taken on Nikon Ti-2E Inverted Microscope. GFP is shown with DAPI overlay; ×20 objective with 1.5 magnification; scale bar, 500 μm. (B) A 30X single region of interest originating from white boxes in images in (A); ×20 objective with 1.5 magnification; scale bar, 50 μm. (C) z stack maximum intensity projection images with orthogonal views taken on Zeiss 800 LSM Confocal with ×20/0.8 M27 objection and 1.5 digital zoom. Representative images show GFP (green) expression with RCVRN + photoreceptors in red. Scale bar, 50 μm. GCL, ganglion cell layer; GFP, green fluorescent protein; INL, inner nuclear layer; ONL, outer nuclear layer; RCVRN, recoverin.

Article Snippet: GFP is shown with DAPI overlay; ×20 objective with 1.5 magnification; scale bar, 500 μm. (B) A 30X single region of interest originating from white boxes in images in (A); ×20 objective with 1.5 magnification; scale bar, 50 μm. (C–G) z stack maximum intensity projection images with orthogonal views taken on Zeiss 800 LSM Confocal microscope with ×20/0.8 M27 objection and 1.5 digital zoom; scale bar, 50 μm.

Techniques: Plasmid Preparation, Injection, Transduction, Inverted Microscopy, Expressing

Journal: Molecular Therapy. Methods & Clinical Development

Article Title: In-depth comparison of Anc80L65 and AAV9 retinal targeting and characterization of cross-reactivity to multiple AAV serotypes in humans

doi: 10.1016/j.omtm.2023.05.016

Figure Lengend Snippet: Anc80 and AAV9 vectors primarily transduce ganglion cells and display different inner nuclear layer tropisms after intravitreal injection in WT C57Bl/6J mice Representative immunofluorescent images of retinas comparing AAV9.CB.GFP (n = 4), Anc80.CB.GFP (n = 3), AAV9.546.GFP (n = 3), and Anc80.546 (n = 3) transduction after intravitreal injection. (A) Large-scan 30X stitched image of whole retina taken on Nikon Ti-2E Inverted Microscope. GFP is shown with DAPI overlay; ×20 objective with 1.5 magnification; scale bar, 500 μm. (B) A 30X single region of interest originating from white boxes in images in (A); ×20 objective with 1.5 magnification; scale bar, 50 μm. (C–G) z stack maximum intensity projection images with orthogonal views taken on Zeiss 800 LSM Confocal microscope with ×20/0.8 M27 objection and 1.5 digital zoom; scale bar, 50 μm. Double immunolabeling of GFP in green with Brn3a + ganglion cells (C), AP-2α + amacrine cells (D), Sox2 + Müller glia (E), Otx2 + bipolar cells (F), and Calb + horizontal cells (G) in red. Representative images for each counterstain within the same treatment group originate from different sections of the same eye or from multiple mice and demonstrate heterogeneous transduction. Calb, calbindin; GCL, ganglion cell layer; GFP, green fluorescent protein; INL, inner nuclear layer; ONL, outer nuclear layer.

Article Snippet: GFP is shown with DAPI overlay; ×20 objective with 1.5 magnification; scale bar, 500 μm. (B) A 30X single region of interest originating from white boxes in images in (A); ×20 objective with 1.5 magnification; scale bar, 50 μm. (C–G) z stack maximum intensity projection images with orthogonal views taken on Zeiss 800 LSM Confocal microscope with ×20/0.8 M27 objection and 1.5 digital zoom; scale bar, 50 μm.

Techniques: Injection, Transduction, Inverted Microscopy, Microscopy, Immunolabeling