Eight mid-lactation Holstein-Friesian dairy cows were used in an experiment to investigate whether poor transfer to milk of (n-3) long chain (≥C20) poly-unsaturated fatty acids (n-3 LCPUFA), from diets containing fish oil, is related to their mode of transport in plasma lipoproteins. The cows were split into two groups of four, each of which was housed with 24 other cows which did not provide blood data in this experiment. All cows received a basal diet ad libitum consisting of (kg/t dry matter) silage (539), grain distillers’ grains (148), rolled barley (248) and soya (65). All cows also received a mid-day meal of 2·76 kg per cow per day of molassed sugar-beet shreds. The control group (C) received no oil supplement but those in the treatment group (F) received 300 g per cow per day of fish oil absorbed onto the sugar-beet shreds. Fish oil inclusion progressively depressed milk fat concentration of the four cows from each group over the 3 weeks of the experiment so that levels were approximately 20 g/kg lower for cows in treatment F than for those in C (P < 0·05). Mean concentrations over all cows of the long-chain (≥C20, n-3) poly-unsaturated fatty acids in milk fat (g/100 g) were 0·12 and 0·54 for treatments C and F respectively, whilst corresponding output in milk (g/day) increased from 116 to 3·91, an average efficiency of transfer of 4·6%. The fatty acid compositions of the lipoproteins were measured in plasma samples from the four cows from each group after 3 weeks on the experimental diets The proportions (g/100 g fatty acids) of C20: 5 (n-3) in cholesteryl (CE) and phospholipid (PL) of high-density lipoprotein (HDL) in the plasma were respectively increased from 0·97 to 4·39 (P < 0·05) and from 1·15 to 4·74 (P < 0·01) by the fish-oil supplementation. For C22: 6 (n-3), the proportions in CE and PL of HDL respectively increased (P < 0·001) from 0·02 to 0·23 and from 0·16 to 2·18 as a result of the supplementation. The mean proportions (g/100 g fatty acids) of C20: 5 (n-3) and C22: 6 (n-3) in the triglyceride fraction of very low-density lipoprotein were 1·05 and 2·3 respectively and were not significantly altered by fish-oil supplementation (P > 0·05). It is suggested that the low transfer efficiency of (n-3) LCPUFA to the milk is partly due to their transport in the plasma largely as components of HDL lipids which are unavailable to the mammary gland.