Objective To establish rabbit models of mixture-infectious endophthalmitis induced by exogenous Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli). Methods A total of 84 eyes of 42 New Zealand white albino rabbits were randomly divided into 4 groups. There were 21 eyes in each group. Rabbit eyes in group 1, 2, 3 and 4 received an intravitreal injection of 0.1 ml of mix bacterium (2times;104 CFU/ ml, including 103 S. aureus and 103 E. coli), S. aureus (104 CFU/ ml), E. coli (104 CFU/ml), and sterilized saline respectively. The eyes were examined by slit-lamp microscopy, ophthalmoscopy, A/B scan, electroretinography (ERG) and bacterial culture of vitreous humors at the timepoints of 6, 12, 24, 48 and 72 hours, and 4, 7, 10, 14 days after intravitreal injection. All eyeballs were then enucleated for histopathological examination. Results Various degrees of inflammatory reactions were presented in the 3 experimental groups after the injection, and the development trend of the disease was nearly the same. In group 1 active intraocular inflammation like anterior chamber exudates, started at 12 hours after injection (which was early than that in group 2 and 3), aggravated between 48 and 72 hours, alleviated slowly from 4 to 7 days, and was obviously better after 10 to 14 days while the corneal neovascularization and vitreous gray opacity begun to form. The bacterial culture was positive in group 1 (100％, 6 hours to 14 days after injection), group 2 (100%, 6 hours to 3 days after injection) and group 3 (100% from 6 hours to 7 days, and 67.67% at 14 days after injection). It was negative for group 2 (7 to 14 days after injection) and group 4 (6 hours to 14 days after injection). The amplitude of ERG b wave dissapeard in group 1 to 3, and decreased less than 30% in group 4 from the 48th hour after injection. Histopathological examination revealed that all intraocular structures infiltrated with inflammatory cells. Conclusion Complicated endophthalmitis rabbit models can be successfully established by intravitreal injection with S. aureus and E. coli.