Mucociliary transport is a major component of respiratory function and contributes greatly to the protection of the airways against infection. Cystic fibrosis is associated with severe abnormalities of the capacities of transport, due to the abnormal mucus and inflammatory changes of the epithelium. Recombinant human DNase I (rhDNase) has been demonstrated to reduce the viscosity of purulent cystic fibrosis respiratory mucus. To investigate the impact of rhDNase on transport capacities, we analysed ex vivo its effect on mucus transport by the ciliary activity and by the simulated cough. We measured in parallel the changes in rheological and physical properties of mucus. In 15 patients with cystic fibrosis, rhDNase was able in vitro to improve mucus transport capacities by simulated cough. The effect was dose-dependent and correlated with a decrease in viscosity and an improvement of surface properties of the mucus. A significant improvement of mucociliary transport was observed only in the subgroup of patients whose transport capacities were altered before incubation with rhDNase. Our findings suggest that rhDNase in vivo might benefit most to patients whose mucus transport capacities are altered, because of highly viscous and tenacious secretions.