Sustained suppression of VEGF is needed in many patients with neovascular age-related macular degeneration (NVAMD), and gene transfer of a VEGF-neutralizing protein is a promising approach to achieve it. Initial clinical trials testing this approach have shown encouraging signals, but evidence of robust transgene expression and consistent antiangiogenic and antipermeability activity has been lacking. In this study, we demonstrate expression of an anti-human VEGF antibody fragment (antiVEGFfab) after subretinal injection of AAV8-antiVEGFfab. In transgenic mice expressing human VEGF in retina (rho/VEGF mice), a model of type 3 choroidal neovascularization (NV), eyes injected with ≥1 × 10⁷ gene copies (GC) of AAV8-antiVEGFfab had significantly less mean area of NV than null vector-injected eyes. A dose-dependent response was observed with modest reduction of NV with ≤3 × 10⁷, >50% reduction with ≥1 × 10⁸ GC and almost complete elimination of NV with 3 × 10⁹ or 1 × 10¹⁰ GC. In Tet/opsin/VEGF mice, in which doxycycline-induced high expression of VEGF leads to severe vascular leakage and exudative retinal detachment (RD), reduction of total RD by 70%–80% occurred with 3 × 10⁹ or 1 × 10¹⁰ GC of AAV8-antiVEGFfab, an effect that was sustained for at least a month. These data strongly support initiating clinical trials testing subretinal injection of AAV8-antiVEGFfab in patients with NVAMD.