We have previously demonstrated that recombinant human Cu/Zn superoxide dismutase (rhCu/ZnSOD) is rapidly incorporated into cells of airways, respiratory bronchioles and alveoli following intratracheal (IT) administration. The present study examined whether this cellular uptake is specific to Cu/Zn SOD, or if other proteins are similarly incorporated into lung cells. Two different proteins were used, rhMnSOD and human albumin, which were labeled with the fluorescent dye SFX prior to IT administration. Fourteen newborn piglets (2 to 3 days old, 1.2 to 2.0 kg) were intubated and mechanically ventilated. Four piglets received rhMnSOD and 6 received albumin. Protein was delivered with the animals' right side down and head elevated to enhance deposition in the right lower lobe (RLL). In addition, to determine whether endogenous surfactant was necessary for intracellular localization, four piglets were made surfactant deficient by repeated bronchoalveolar lavage prior to protein delivery. All animals were sacrificed at 30 or 60 minutes after instillation. The lungs were fixed by perfusion in buffered paraformaldehyde, removed and cryosections of the RLL were examined using laser confocal microscopy in a blinded fashion. All images were acquired under identical conditions. Similar to our previous observations with rhCu/ZnSOD, intracellular uptake of larger molecular weight proteins, rhMnSOD and albumin, was noted throughout the lung. This uptake was not affected by surfactant deficiency. We conclude that the cellular uptake of protein delivered by the IT route occurs via a nonspecific mechanism. We speculate that this mechanism exists to clear proteins that accumulate in the lumen of the airways and the alveoli. Endogenous surfactant does not appear to have a role in the uptake process. (Funded by grants from the American Lung Association and Winthrop-University Hospital)