The development and performance of a novel nanoparticle-based formulation for pulmonary delivery has been characterized chronologically through the particle preparation process, in vitro testing of drug release, biocompatibility, degradation, drug transport in cell culture, and in vivo bronchoprotection studies in anaesthetised guinea pigs. This study demonstrates excellent agreement of the in vitro and in vivo experiments undertaken to prove the feasibility of the design, thereby serving as an example highlighting the importance of in vitro test methods that predict in vivo performance. Nanoparticles were prepared from the newly designed negatively-charged polymer poly(vinyl sulfonate-co-vinyl alcohol)-g-poly(d,l-lactic-co-glycolic acid) loaded with salbutamol free base. Average particle sizes of blank and drug-loaded nanoparticles prepared at the various stages of the investigations were between 91 and 204nm; average zeta potential values were between -50.1 and -25.6mV. Blank nanoparticles showed no significant toxicity, and no inflammatory activity was detected in Calu-3 cells. Sustained release of salbutamol from the nanoparticles was observed for 2.5h in vitro, and a prolonged effect was observed for 120min in vivo. These results demonstrate good agreement between in vitro and in vivo tests and also present a promising foundation for future advancement in nanomedicine strategies for pulmonary drug delivery.