Fish oil (FO) addition to dairy cow diet decreases the milk fat content. This could facilitate the management of milk fat quotas by farmers, but could also change the milk quality. The aim of this study was to compare the respective effects of a control diet (C), and either a ruminal (R) or a duodenal (D) infusion of FO (menhaden type, 300mL/day for 4 weeks), on milk yield and composition (fat and protein contents, fatty acid profile). A 3 3 latin square was designed on 6 mid-lactation Holstein cows fitted with rumen and duodenum cannulas.FO infusions did not change milk yield (22.7 kg/day) despite decreases in oil-free dry matter intake (C, 19.8; D, 17.7; R, 15.9 kg/day). Milk protein content was significantly lower with R infusion (C, 29.7; D, 28.6; R, 27.8 g/kg), and milk fat content with both infusions (C, 35.4; D, 32.2; R, 25.1 g/kg).The decrease in milk fat yield (-46 g/day) with D infusion was mainly due to a decrease in C16:0 (-52 g/day) and cis-C18:1 (-13 g/day) secretion, that was compensated in part by an increase in C20:5 (+10 g/day) and C22:6 (+3 g/day) secretion. The sharp decrease in milk fat yield (-216 g/day) with R infusion was due to a lower secretion of C4:0 to C14:0 (-67 g/day), C16:0 (-79 g/day), C18:0 (-45 g/day) and cis-C18:1 (-98 g/day), that was compensated in part by a higher secretion of trans-C18:1 (+63 g/day), C20:5 (+1 g/day), C22:6 (+2 g/day) and other C20-C22 fatty acids (+28 g/day).Unprotected FO addition to dairy cow diet (R infusion) was shown to decrease the milk protein content, to decrease sharply the milk fat content and to increase the secretion of trans-C18:1, that arose from changes in rumen ecosystem and was probably the cause of the collapse of mammary lipogenesis. Protected FO addition (D infusion) would decrease the milk fat content and increase the n-3 polyunsaturated fatty acid secretion.