Helmholtz Institute for Pharmaceutical Research Saarland (HIPS) 1 article published in JoVE Immunology and Infection P. aeruginosa Infected 3D Co-Culture of Bronchial Epithelial Cells and Macrophages at Air-Liquid Interface for Preclinical Evaluation of Anti-Infectives Carlos Victor Montefusco-Pereira*1,2, Justus C. Horstmann*1,2, Thomas Ebensen3, Christoph Beisswenger4, Robert Bals4, Carlos A. Guzmán3, Nicole Schneider-Daum1, Cristiane de Souza Carvalho-Wodarz1, Claus-Michael Lehr1,2 1Department of Drug Delivery, Helmholtz Institute for Pharmaceutical Research Saarland (HIPS), 2Department of Pharmacy, Saarland University, 3Department of Vaccinology and Applied Microbiology, Helmholtz Centre for Infection Research, 4Department of Internal Medicine V - Pulmonology, Allergology, Respiratory Intensive Care Medicine, Saarland University Hospital We describe a protocol for a three-dimensional co-culture model of infected airways, using CFBE41o- cells, THP-1 macrophages, and Pseudomonas aeruginosa, established at the air-liquid interface. This model provides a new platform to simultaneously test antibiotic efficacy, epithelial barrier function, and inflammatory markers.