Three dry powder formulations of tiotropium bromide were manufactured for in vitro comparison to Spiriva® HandiHaler®. In this study, aerodynamic particle size distribution (APSD) characterization techniques that simulate patient conditions coupled with particle chemistry, morphology and size analysis were used to determine differences in the tiotropium bromide formulations. The objective of this study was to determine the comparability of the formulations using traditional and novel in vitro methods.