Substantial advances in our understanding of lentivirus lifecycles and their various constituent proteins have permitted the bioengineering of lentiviral vectors now considered safe enough for clinical trials for both lethal and non-lethal diseases. They possess distinct properties that make them particularly suitable for gene delivery in ophthalmic diseases, including high expression, consistent targeting of various post-mitotic ocular cells in vivo and a paucity of associated intraocular inflammation, all contributing to their ability to mediate efficient and stable intraocular gene transfer. In this review, the intraocular tropisms and therapeutic applications of both primate and non-primate lentiviral vectors, and how the unique features of the eye influence these, are discussed. The feasibility of therapeutic targeting using these vectors in animal models of both anterior and posterior ophthalmic disorders has been established, and has, in combination with substantial progress in enhancing lentiviral vector bio-safety over the past two decades, paved the way for the first human ophthalmic clinical trials using lentivirus-based gene transfer vectors.