Journal: Investigative Ophthalmology & Visual Science

Article Title: AAV2-Retro–Mediated Gene Transfer Selectively Targets Outer Retinal Cells Following Intravitreal Injection

doi: 10.1167/iovs.67.3.54

Figure Lengend Snippet: AAV2-retro preferentially transduces photoreceptors and RPE 1 and 3 days after intravitreal injection. ( A , C ) Stitched 10× images of retinal cross-sections showing mGL expression at 1 dpi ( A ) and at 3 dpi ( C ). Blue indicates DAPI-labeled nuclei; green indicates mGL reporter expression. ( B , D ) Higher-magnification images of representative retinal regions showing mGL expression at 1 dpi ( B ) and at 3 dpi ( D ). ( B’ , D’ ) Merged images corresponding to panels B and D , illustrating mGL and DAPI expression across retinal layers. Retinal layers are indicated: RPE, ONL, OPL, INL, inner plexiform layer (IPL), and GCL. Scale bars : 200 µm ( A , C ); 50 µm ( B’ , D’ ).

Article Snippet: AAVs were produced with two different capsids: AAV2-retro (rAAV2-retro helper was a gift from Alla Karpova and David Schaffer; Addgene plasmid #81070; RRID:Addgene_81070) and MNM008.

Techniques: Injection, Expressing, Labeling

Journal: Investigative Ophthalmology & Visual Science

Article Title: AAV2-Retro–Mediated Gene Transfer Selectively Targets Outer Retinal Cells Following Intravitreal Injection

doi: 10.1167/iovs.67.3.54

Figure Lengend Snippet: AAV2-retro maintains selective transduction in the outer layers 14 days after intravitreal injection. ( A ) Stitched 10× images of retinal cross-sections showing mGL expression at 14 dpi. ( B ) Higher-magnification view of a representative retinal region with mGL expression. ( B’ ) Merged image from B highlighting layer-specific distribution (RPE, ONL, OPL, INL, IPL, and GCL), mGL, and DAPI across the layers. ( C ) Retinal flatmount at 14 dpi after sequential injections, showing widespread mGL expression. ( C’ , C’’ ) Higher magnification images of the boxed areas in C . Scale bars : 200 µm ( A , C ); 50 µm ( B’ , C’ , C’’ ). ( D ) Quantification of mGL-positive cells within the regions of high transduction in the ONL (i.e., ROI). The percentage shown indicates the proportion of transduced cells among total DAPI-positive ONL cells. N = 3 animals. Scale bars : 200 µm ( A , C ); 50 µm ( C’ , C’’ ).

Article Snippet: AAVs were produced with two different capsids: AAV2-retro (rAAV2-retro helper was a gift from Alla Karpova and David Schaffer; Addgene plasmid #81070; RRID:Addgene_81070) and MNM008.

Techniques: Transduction, Injection, Expressing

Journal: Investigative Ophthalmology & Visual Science

Article Title: AAV2-Retro–Mediated Gene Transfer Selectively Targets Outer Retinal Cells Following Intravitreal Injection

doi: 10.1167/iovs.67.3.54

Figure Lengend Snippet: AAV2-retro leads to similar preferential transduction in female mice after intravitreal injection. ( A , C ) Stitched 10× images of retinal cross-sections showing mGL expression at 3 dpi ( A ) and at 14 dpi ( C ). Blue indicates DAPI-labeled nuclei; green indicates mGL reporter expression. ( B , D ) Higher magnification images of representative retinal regions showing mGL expression at 3 dpi ( B ) and at 14 dpi ( D ). ( B’ , D’ ) Merged images corresponding to panels B and D , illustrating mGL and DAPI expression across retinal layers. Retinal layers are indicated: RPE, ONL, OPL, INL, IPL, and GCL. Three female mice were examined per time point with similar results. Scale bars : 200 µm ( A , C ); 50 µm ( B’ , D’ ).

Article Snippet: AAVs were produced with two different capsids: AAV2-retro (rAAV2-retro helper was a gift from Alla Karpova and David Schaffer; Addgene plasmid #81070; RRID:Addgene_81070) and MNM008.

Techniques: Transduction, Injection, Expressing, Labeling

Journal: Investigative Ophthalmology & Visual Science

Article Title: AAV2-Retro–Mediated Gene Transfer Selectively Targets Outer Retinal Cells Following Intravitreal Injection

doi: 10.1167/iovs.67.3.54

Figure Lengend Snippet: AAV2-retro effectively delivers transgene to both rods and cones. ( A , B ) Representative retinal cross-sections showing mGL expression ( green ) at 3 dpi ( A ) and at 14 dpi ( B ) following intravitreal injection of AAV2-retro–CMV–mGL. Cone arrestin ( magenta ) was used to identify cone photoreceptors. Bottom panel in A shows higher magnification images highlighting the colocalization of mGL with cone arrestin immunoreactivity. Scale bars : 50 µm ( A , B ); 5 µm (higher magnification images).

Article Snippet: AAVs were produced with two different capsids: AAV2-retro (rAAV2-retro helper was a gift from Alla Karpova and David Schaffer; Addgene plasmid #81070; RRID:Addgene_81070) and MNM008.

Techniques: Expressing, Injection

Journal: Investigative Ophthalmology & Visual Science

Article Title: AAV2-Retro–Mediated Gene Transfer Selectively Targets Outer Retinal Cells Following Intravitreal Injection

doi: 10.1167/iovs.67.3.54

Figure Lengend Snippet: AAV2-retro transduces only a small number of amacrine cells and RGCs. ( A , B ) Retinal cross-sections show mGL expression ( green ) at 3 dpi ( A ) and at 14 dpi ( B ). AP-2α immunoreactivity is shown in magenta and RBPMS immunoreactivity in red . (A’ , B’ ) Higher magnification images showing cells that appear to be co-labeled with mGL and AP-2α. ( C ) Quantification shows that the percentage of mGL + cells that were positive for either RBPMS (in GCL and INL) or AP-2α at each time point. N = 3 per group. Error bar, ± SEM Two-way ANOVA with Tukey's multiple comparisons: * < 0.05, **** < 0.0001, ns = not significant. Scale bars : 50 µm ( A , B ); 5 µm ( A’ , B’ ).

Article Snippet: AAVs were produced with two different capsids: AAV2-retro (rAAV2-retro helper was a gift from Alla Karpova and David Schaffer; Addgene plasmid #81070; RRID:Addgene_81070) and MNM008.

Techniques: Expressing, Labeling