Reactive oxygen species (ROS) are formed under physiological and pathophysiological conditions. High-yielding dairy cows exhibit a high metabolic activity during lactation that increases the risk of oxidative stress (OST). Fatty acids (FA) can increase ROS but specific essential FA and conjugated linoleic acid (CLA) are also discussed to prevent OST. Common diets in dairy farming often deliver lower amounts of n-3 FA compared with pasture based systems. The objective was to evaluate the effects of a diet low in n-3 FA content, followed by FA supplementation on the oxidative status (OS) in red blood cells (RBC). Four rumen fistulated Holstein cows (126 ± 4 d in milk) fitted with abomasal infusion tubes were arranged in a 4 × 4 Latin square model. Cows were fed a TMR based on corn silage and supplemented twice per day with 3 successively rising lipid dosages. Each dosage was given for 2 wk, resulting in a 6-wk treatment period, followed by a 3-wk wash out phase. Supplements were coconut oil delivering medium-chain FA (38, 77 and 153 g/d, respectively), linseed-safflower oil mix, delivering n-3 FA (EFA; 42, 82 and 163 g/d, respectively), LUTALIN (c9,t11 and t10,c12 CLA; BASF SE, Ludwigshafen, Germany, in equal amounts; 16, 32 and 64 g/d, respectively), or EFA + CLA. Milk and blood plasma were sampled every 2 weeks. The RBC were isolated at the end of the highest dosage of each treatment and wash out period. Indicators of OST as superoxide dismutase, glutathione peroxidase, malondialdehyde and reactive oxygen metabolites (d-ROMS) were measured. Data were analyzed by mixed model of SAS using repeated measurement analysis with treatment as fixed effect and week in milk as covariate. LSMEANS were compared by Tukey test. Linolenic acid as well as c9,t11 and t10,c12 CLA increased dose dependently (P < 0.05) in milk fat and plasma of the respective treatment groups. The analyzed indicators of OST in RBC were not affected by the different supplements. These data indicate that a diet low in n-3 FA may has no effects on the OS in RBC of mid-lactating dairy cows.