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Objective: To develop a protocol for the treatment of intraocular lymphoma by the intravitreal injection of methotrexate. Methods: Patients whose results were negative for human immunodeficiency virus and who had pathologically confirmed intraocular lymphoma were eligible for participation in the study. A dose of 400 pg of methotrexate was given intravitreally twice weekly until the vitreous was clinically cleared of cells. Weekly injections were then given for 1 month, followed by monthly injections for 1 year. Results: Seven eyes of 4 patients were treated. Three patients have completed the protocol. To date, only 1 eye has suffered a significant loss of vision. No serious ocular toxic reaction has been identified. Conclusions: Survival rates and time to relapse for patients with primary central nervous system lymphoma have improved. The role of ocular radiation therapy must be weighed against the potential drawbacks. Injecting chemotherapeutic agents into the vitreous is worthy of consideration. Four patients treated to date at our institution have had promising results. Intravitreal chemotherapy may result in improved treatment of intraocular lymphoma with reduced morbidity.