Ventilation in cities is crucial for thewell being of their inhabitants. Therefore, local governments require air ventilation assessments (AVAs) prior to the construction of new buildings. In a standard AVA, however, only neutral stratification is considered, although diabatic and particularly unstable conditions may be observed more frequently in nature. The results presented here indicate significant changes in ventilation within most of the area of Kowloon City, Hong Kong, included in the study. A new definition for calculating ventilation was introduced, and used to compare the influence of buildings on ventilation under conditions of neutral and unstable stratification. The overall ventilation increased due to enhanced vertical mixing. In the vicinity of exposed buildings, however, ventilation was weaker for unstable stratification than for neutral stratification. The influence on ventilation by building parameters, such as the plan area index, was altered when unstable stratification was considered. Consequently, differences in stratification were shown to have marked effects on ventilation estimates, which should be taken into consideration in future AVAs.