This study compared the effects of two levels of crude protein (CP) fed during late gestation on the performance, blood metabolites, and ovarian activity of Holstein cows. One-hundred and six cows (42 primigravid and 64 multiparous) 32 d before calving were divided into two groups and fed diets containing moderate (12.7% CP, 36% rumen undegradable protein, (RUP) or high (14.7% CP, 40% RUP) protein. Higher prepartum CP diet increased milk production during the first 120 d in milk (DIM), but most of that effect was detected for the primigravid cows. Primigravid cows fed the prepartum diet higher in protein produced 2.0 kg/d more milk and 3.1 kg/d more 3.5% fat-corrected milk (FCM) during early lactation. Yields of milk fat and protein in early lactation were also increased by the high prepartum CP diet fed to primigravid cows. During the complete lactation, the response to prepartum dietary protein differed between primigravid and multiparous cows. Yields of milk, 3.5% FCM, and milk fat and protein were not affected by the prepartum diet for primigravid cows, but decreased for multiparous cows fed the high protein diet. However, primigravid cows fed the high prepartum protein diet had a higher 305-d mature equivalent milk yield. Colostrum composition, blood metabolites, ovarian activity, and disease incidence were not influenced by prepartum protein. Data from this study suggest that the current prepartum protein recommendation seems to be adequate for multiparous cows, but late-gestation primigravid cows might benefit from diets with a CP content above 12.7%.