Thirty mink dams nursing litters of six kits were assigned to one of three dietary treatments [high protein (HP), medium protein (MP) and low protein (LP)], fed ad libitum for 4 week from parturition, to investigate the effects of protein supply on milk yield and milk composition in order to estimate the amino acid requirement of the lactating mink. Twelve dams were held in an intensive care unit and subjected to balance experiments and the kits were injected with deuterium oxide to determine water kinetics and milk yield. Eighteen dams were kept under normal farm conditions but with feed intake of dams and live weight gain of kits being determined and milk samples collected. The ME intake was higher (p < 0.05) in dams fed the LP and MP diets than in dams fed the HP diet, whereas the amino acid intake (g/day) was lowest (p < 0.05) in dams fed the LP diet. In the third and fourth weeks of lactation milk yield was higher (p < 0.05) in dams fed the LP and MP diets than in dams fed the HP diet. Chemical composition of milk was not affected (p > 0.05) by dietary treatment. However, protein content tended (p = 0.06) to be lower in dams fed the LP diet. Amino acid content (g/16 g N) of milk was higher (p < 0.05) in dams fed the LP and MP diets than in dams fed the HP diet. This resulted in the highest (p < 0.05) amino acid intake and highest (p < 0.001) live weights of kits nursed by dams fed the LP and MP diets, which may be explained by a combined effect of higher ME intake and reduced energetic costs for glucose production through less amino acids being used in gluconeogenesis. In conclusion, the improved performance of dams fed the LP diet suggested that their requirement of essential amino acids and non-specific N were covered, and the requirement of digestible amino acids of lactating mink (kg(0.75)) was, thereby, estimated by use of a factorial approach including the amino acid excretion in milk of LP dams.