The objective of this study was to evaluate the effects of supplementation of dairy cows with different fatty acid sources (soybean oil (SO) and fish oil (FO)) on milk production, milk composition, milk fatty acid profile, and physicochemical and sensory characteristics of ice cream. During 63 days, fifteen Holstein cows averaging 198 ± 35 days in milk were assigned to three groups: control diet with no added lipid (n = 5 cows); and supplemented diets with SO (n = 5 cows; unrefined SO; 30 g/kg DM) or FO (n = 5 cows; FO from unrefined salmon oil; 30 g/kg DM). Milk production, milk fat, and milk protein were not affected by treatments. Saturated fatty acids in milk fat were decreased with SO and FO compared with control. C18:2 cis-9, cis-12 was increased with SO whereas C18:2 cis-9, trans-11, C20:3n-3, C20:3n-6, C20:5n-3, and C22:6n-3 were the highest with FO. Draw temperature and firmness were higher in SO compared to control and FO ice creams. Melting resistance was higher in FO compared with control and SO ice creams. Supplementation of cow diets with SO and FO did not have detrimental effects on milk production, or ice cream physicochemical and sensory characteristics.