Three hundred one-day-old avian broilers were divided into four equal groups of 75 animals that were fed for 42 days as follows: a control diet containing 23 mg fluorine (F)/kg and three high F diets containing 400, 800, and 1200 mg F/kg, respectively, for high F groups I, II, and III. The superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were greatly decreased, while the malondialdehyde (MDA) contents were markedly increased in high F groups II and III. At the same time, mitochondrial injury and expanded endocytoplasmic reticulum were obviously observed in high F groups II and III, and the fluoride contents both in spleen and serum were significantly increased in the three high F groups when compared with those of control group. The results showed that excess dietary F in the range of 800−1200 mg/kg caused obvious oxidative stress, which provided a possible pathway for the apoptosis of splenocytes in chickens.