Ocular infections may arise spontaneously or following penetrating globe injury or operation, such as corneal transplant or cataract extraction. Treatment and prophylaxis of bacterial infections using antibiotic eye drops requires a strict dosing regimen to avoid irreversible vision loss. At present, moxifloxacin eye drops are prescribed for use multiple times per day, leading to patient non-adherence, the emergence of bacterial resistance, and infection progression. The desire to avoid sub-lethal antibiotic dosing and visual impairment through inconsistent eye drop application motivates the development of a sustained release injectable formulation. Herein, we report the development of an ion-paired, nanocrystalline moxifloxacin formulation that provided increased intraocular antibiotic accumulation with a single subconjunctival injection compared to 3× daily eye drops. The sustained release functionality further led to improved or non-inferior prevention and treatment of Staphylococcus aureus-induced ocular infection in both rats and rabbits compared to gold standard intracameral moxifloxacin and moxifloxacin eye drops. By achieving therapeutically relevant moxifloxacin accumulation, this nanocrystalline moxifloxacin formulation may be a promising alternative to conventional therapies to achieve improved post-surgical infection prevention and bacterial keratitis treatment outcomes.