Using inhalable nanoparticles (NPs) to treat pulmonary diseases is an emerging field (1). Lungs as part of the mononuclear phagocyte system (MPS) are extensively rich with alveolar macrophages (2). These alveolar macrophages are part of the body’s immune system and responsible for clearing inhaled foreign objects from the alveolar spaces (3), which cause a substantial portion of the inhaled NPs to be cleared after inhalation once NPs are settled in the alveolar spaces (4). Clearance of NPs by macrophages is not exclusive to inhalable NPs; as it is associated with other routes of administrations, such as intravenous i.v. (5). In fact, the failure of early attempts of using NPs for cancer targeting was mainly attributed to the interference of macrophages of the MPS. The early work showed that after i.v. injection, NPs were cleared immediately by phagocytotic cells, specifically by macrophages and Kuppfer cells of the liver (6). Thus, in order to evade macrophages and increase NPs circulation, a new strategy was implemented by changing the NPs surface properties (5). This was accomplished by using hydrophilic surfactants, such as polysorbate 80 to coat the NPs surface (7). Hydrophilic coating of NPs changes the physicochemical characteristics of NPs. This inhibits opsonization, which is the precipitation of antibodies and other proteins on the surface of NPs (8). This strategy was shown to successfully prolong the circulation time of NPs and also enhanced the distribution of NPs to some organs such as the brain (9). Using coated NPs for pulmonary delivery may be associated with increased bioavailability since inhaled NPs may have a higher chance to evade alveolar macrophages. However, the delicate composition of the alveolar space should stay unchanged and kept intact after delivery of any inhalable treatment including NPs. In the alveolar space, a thin film of lung surfactant covers the epithelium. It is composed of phospholipids and proteins (10). The main component of the film is 1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine (DPPC), a phosphatidylcholine with two saturated 16-carbon long fatty acyl chains (11). This phospholipid is a key factor for controlling the surface tension and the gas exchange process in the alveolar sacs (12). Moreover, the ability of the lungs to function properly depends on the integrity of this surfactant layer (13). In vitro, lung-surfactant model systems have been developed to study the impact of NPs on the surfactant layer (14). Changes in the surface pressure-area isotherms induced by NPs were recorded and considered as an indicator for interactions of NPs and components of the surfactant model. Previous studies investigated the effect of non-coated NPs on the integrity of the lung surfactant film in vitro (10,11,14). Azarmi et al. showed in vivo that non-coated NPs are well tolerated by Balb/c nude mice after pulmonary administration (15). However, the impact of coated NPs on the integrity of the surfactant film is yet unknown. Therefore, the aim of this study was to investigate the biophysical interactions of polysorbate-80-coated NPs with DPPC monolayer film in vitro. The results were compared with in vivo observations.