Neutrophil elastase (NE) contributes to progression of the lung disease characteristic of cystic fibrosis (CF). We developed a strategy that permits the delivery of α₁-antitrypsin ( α₁-AT) to inaccessible CF airways by targeting the respiratory epithelium via the polymeric immunoglobulin receptor (pIgR). A fusion protein consisting of a single-chain Fv directed against human secretory component (SC) and linked to human α₁-AT was effectively transported in a basolateral-to-apical direction across in vitro model systems of polarized respiratory epithelium consisting of 16HBEo cells transfected with human pIgR complementary DNA, which overexpress the receptor, and human respiratory epithelial cells grown in primary culture at an air–liquid interface. When applied to the basolateral surface, the anti-SC Fv/ α₁-AT fusion protein penetrated the respiratory epithelia, with transcytosis of the fusion protein being related to the amount of SC detected at the apical surface. Significantly less fusion protein crossed the cells in the opposite direction. In addition, because the antihuman SC Fv/ α₁-AT fusion protein was transported vectorially and deposited into the small volume of apical surface fluid, the antiprotease component of this protein was concentrated atop the epithelium. Thus, in cell models, this system is capable of concentrating the antiprotease of the fusion protein, in the thin film of epithelial surface fluid to a level expected to be therapeutic in the airways of many patients with CF. Ferkol T, Eckman E, Swaidani S, Silski C, Davis P. Transport of bifunctional proteins across respiratory epithelial cells via the polymeric immunoglobulin receptor.