Essential amino acids most limiting for milk production and/or secretion of milk protein were determined in five trials involving infusion of amino acids into the abomasum of lactating Holstein cows. Cows were fed corn-based rations containing 10.7 to 11.5% crude protein (dry matter basis) which exceeded standard allowances for energy. All trials were Latin-square designs with 9-day periods. The abomasal infusion of methionine had no effect on secretion of milk, milk protein, or milk fat. Lysine infusion resulted in 16% of the total response in yield of milk protein that was obtained with either the 10 essential amino acids or sodium caseinate while infusion of lysine and methionine together accounted for 43% of the total response. This suggested that lysine and methionine were first and second limiting, or co-limiting, for secretion of milk protein when rations consisting primarily of corn, corn silage, and alfalfa-grass hay were fed. The primary effect was on content of milk protein rather than milk yield. In general, amino acid infusions had no effect on feed intake, milk fat, and nonprotein nitrogen content of milk. Little change occurred in concentrations in plasma of other amino acids when lysine and methionine were infused together. When concentrations were lowered, indicating increased protein synthesis, changes were too small to be statistically significant.