Understanding mechanisms and quantitative description of cellular responses to changing nutrient supply is critical for predicting milk component yields. Our objective was to evaluate cellular uptake and kinetic behavior of individual amino acids (AA) simultaneously. Bovine mammary epithelial cells were grown to confluency and fed media with an AA profile and concentration similar to dairy cows for 24 h. Treatments were 4 AA concentrations, 0.36, 2.30, 4.28, and 6.24 mM, which represents 16, 100, 186, and 271% of typical plasma AA concentrations in dairy cows. Twenty-four plates of cells (100 × 20 mm) were assigned to each treatment. Cells were first subjected to treatment media enriched with ¹⁵N labeled AA for 24 h and then incubated with treatment media enriched with ¹³C labeled AA for 0, 15, 60, 300, 900, 1800, and 3600 s. Intracellular free AA, intracellular protein-bound AA, and extracellular media AA were analyzed for concentrations and isotopic enrichment using gas chromatography mass spectrometry. Additionally, the total protein, weight, and count of cells were measured. A dynamic 9-pool model was constructed representing extracellular, intracellular, and turnover pools for individual AA and respective isotopes and then solved via numerical integration. Fluxes were extracellular to intracellular entry, intracellular efflux, protein turnover, transamination, oxidation, and synthesis. As an example, cellular influx and efflux ranged from 0.9 to 11.1 and 0 to 9.5 nmol/min for Leu whereas Ala ranged from 1.6 to 50.0 and 0.9 to 56.9 nmol/min for influx and efflux. The average root mean square prediction error as percent of the mean for the 9 pools was 14.6 and 10.6 percent for Leu and Ala respectively. When AA influx was standardized to total pool of the respective AA, Michaelis-Menten kinetics were observed with a K_m of 72 and 345 μmol for Leu and Ala respectively. The described model provides insight on individual AA transport kinetics when subjected to varying extracellular concentrations, which might be useful for better understanding and future modeling of AA uptake by the udder.