The effect of protein intake during gestation on mammary development was evaluated using 17 Landrace x Yorkshire gilts. On d 25 of gestation, gilts were allocated to dietary treatments consisting of levels of protein to achieve lysine intakes of approximately 4, 8, or 16 g/d. Diets provided similar amounts of ME (6.5 Mcal/d) and all other nutrients. On d 105 of gestation, gilts were bled through venipuncture to collect blood samples for analyses of blood metabolites (glucose, NEFA, blood urea nitrogen [BUN]), and metabolic hormones (insulin and IGF-I). Gilts were slaughtered between d 105 and 110 of gestation for complete collection of mammary glands. All mammary glands were carefully removed and separated from each other along the midline. One side was weighed, immediately frozen in liquid nitrogen, and stored at -70°C until further processing. Frozen glands were sliced, and parenchymal tissue was separated and pooled across glands within sow, homogenized, and analyzed for DNA, RNA, and protein concentrations as indices of mammary development. Results showed no effect (P > .10) of level of lysine intake on amount of mammary parenchymal tissue or on concentration or total amount of DNA, RNA, and protein. This experiment did not detect differences between treatments in mammary development as measured by nucleic acids and protein. Therefore, there was no effect of the amount of protein intake by gilts on mammary development. Regression analyses failed to detect any relationships between measures of mammary development and levels of metabolites or metabolic hormones.