Lactoferrin is an iron-binding glycoprotein abundantly present in human milk, and has been postulated both to increase and to decrease intestinal iron absorption. To examine this problem, the interaction of milk iron with pup hemoglobin was studied in controls and in transgenic mice overexpressing human lactoferrin in their milk (2 lines expressing 12 mg/mL and 4 mg/mL, respectively). At day 14 of gestation, pregnant mice were switched from a diet of commercial chow containing iron at 300 mg/kg to diets containing 5, 15, or 50mg iron/kg; controls continued on chow. Nontransgenic pups were cross-fostered to transgenic dams to ensure that any results found in the pups were the effect of milk components. The hemoglobin level in the blood of 10-day-old suckling neonates was measured and calculated as total hemoglobin per pup. The total hemoglobin levels were lower in the pups receiving milk high in human lactoferrin, but the difference reached significance (P < 0.02) only at the highest level of dietary iron. Our findings do not support the hypothesis that lactoferrin functions as an intestinal iron scavenger, at least at high doses.