Four lactating Holstein cows fitted with a rumen cannula and T-type cannulae in the duodenum and ileum were used in an experiment with a 4 X 4 Latin square arrangement of treatments. Diets containing soybean meal (SBM), corn gluten meal (CGM), wet brewers grains (WBG) and distillers dried grains with solubles (DDG) were compared with regard to protein degradability in the rumen and amino acid flow and absorption in the small intestine. The test protein sources provided about 50% of the dietary protein. Spot samples of digesta were collected during a 96-h period and lanthanum (La) was used as an indigestible marker to estimate flow and digestibility of nutrients. Apparent organic matter digestibility in the rumen was 31 +/- 4.8%, while true digestibility based on measurements of microbial organic matter averaged 55 +/- 5.1%. Ruminal ammonia-nitrogen (NH3-N) concentrations (mg/100 ml) were 10.1, 5.6, 6.9 and 6.4 for the SBM, CGM, WBG and DDG diets, respectively. Using diaminopimelic acid as a microbial marker, protein degradation in the rumen was higher for the SBM diet (70%) than for the CGM (45%), WBG (52%) and DDG (46%) diets. Flow of amino acids to the small intestine was generally higher than amino acid intake for all diets. Apparent absorption of amino acids from the small intestine was 70, 77, 71 and 66% of the amino acids entering the duodenum for the SBM, CGM, WBG and DDG diets, respectively. Actual amounts absorbed (g/d) were lowest for the SBM diet. It was concluded that diets containing CGM, WBG or DDG will generally supply more total amino acids to the intestine than a diet containing SBM. Net amounts of amino acids available for absorption in the intestine were also higher for these diets because protein availability in the intestine was not impaired even though protein was more resistant to microbial breakdown in the reticulorumen.