Six lactating Holstein cows, fitted with T-type cannulas in the proximal duodenum, were used in a replicated 3 x 3 Latin square design to determine influence of supplemental protein on amino acid profiles of duodenal chyme and plasma. Protein sources were blood meal, corn gluten meal, and cottonseed meal, which furnished approximately 46% of the total protein in corn-grain corn silage diets. Markers were 15N to estimate rumen bacteria and chromic oxide to estimate nutrient flow. Dry matter intake was lowest on blood meal. Duodenal flow of N exceeded N intake 23% on blood meal and corn gluten meal. Percentages of organic matter and protein digested in the rumen were 56.5, 41.8; 61.2, 53.4; and 50.6, 56.2 for the respective diets. Variation in patterns of amino acids flowing to the duodenum, but not in coccygeal blood, closely reflected dietary differences with six of eight essential amino acids highest for the same treatment in both diet and duodenal chyme. Essential amino acids in least abundance for milk production, measured by mammary extraction coefficients, differed among diets. We conclude that supplemental protein source influences greatly the quantity and pattern of amino acids available for digestion in lactating dairy cows.