The expression of human lactoferrin in the mammary gland is an attractive approach to diminish its current production cost. Previous attempts to produce lactorferrin in the milk of transgenic animals resulted in very high cost and uncertain results. In this paper, we have directly infused replication-defective adenovirus encoding human lactoferrin cDNA into the mammary gland of goats via the teat canal. In this way, we obtained a high level of expressed human lactoferrin up to 2g/L in the milk of goats. The milk serum was collected from the infected mammary gland 48 h post-infection and subjected to a 10% SDS-PAGE and Western blotting. A approximately 80-kDa protein was visualized after viral vector infection. Our results demonstrate that intraductal injection of recombinant replication-defective adenovirus vectors may provide a very useful tool for large-scale production of recombinant proteins of biopharmaceutical interest.