The main objective of this experiment was to examine the effects of the percentage and source of crude protein (CP) and the amount of starch in the diet of dairy cows on the lactational performance and use of N for milk production. Sixty multiparous Holstein cows were used in a 210-d lactational trial with a completely randomized design with a 2 x 3 factorial arrangement of treatments. Two sources of CP [solvent-extracted soybean meal (SBM) and a mixture of SBM and a blend of animal-marine protein supplements plus ruminally protected Met (AMB)] and 3 levels of dietary CP (means = 14.8, 16.8, and 18.7%) were combined into 6 treatments. On a dry matter (DM) basis, diets contained 25.0% corn silage, 20.0% alfalfa silage, 10.0% cottonseed, 26.7 to 37.0% corn grain, and 4.8 to 13.5% protein supplement, plus minerals and vitamins. Across the 210 d of lactation, the productive response of dairy cows to the source of supplemental CP depended on the concentration of CP in the diet. At 18.7% CP, cows fed SBM consumed more DM and produced more milk, 3.5% fat-corrected milk, fat, and true protein, but had lower efficiency of feed use and body condition score than cows fed AMB. At 16.8% CP, cows fed AMB produced more 3.5% fat-corrected milk, fat, and true protein than cows fed SBM. At 14.8% CP, cows fed SBM consumed more DM but produced less true protein and had lower feed efficiency than cows fed AMB. Across CP sources, cows fed 14.8% CP produced less fat-corrected milk and true protein than cows fed 16.8 and 18.7% CP. Across CP percentages, cows fed AMB produced more fat-corrected milk per kilogram of DM consumed than cows fed SBM. Despite these interactions, improvements in the gross efficiency of N use for milk production were achieved through reductions in the intake of N independently of the source of CP. Data suggest that the intake of N by high-producing dairy cows that consume sufficient energy and other nutrients to meet their requirements can be decreased to about 600 to 650 g daily if the source of RDP and RUP are properly matched with the source and amount of carbohydrate in the diet.