The walls in a building envelope have the largest contact area with the exterior environment, and, therefore, a considerable portion of the thermal energy can be lost through the walls compared to the other parts of the building envelope. For energy-saving purposes, the thermal transmittance of walls is typically limited by building energy performance standards at the national level. However, the presence of a ventilated air-space behind the external cladding, which has variable hydro-dynamic behavior, can differently affect the total thermal transmittance of the entire structure. This paper aims to provide an experimental analysis of the total U-value of a ventilated wall assembly measured in a building prototype following the average and dynamic methods defined by ISO 9869-1. Differences between the calculated theoretical U-value and the measured U-value are compared. The contribution of the thermal resistance of the ventilated air-space in the total thermal transmittance of the wall assembly is also analyzed. The results show that the air movement and the enthalpy change in the ventilated cavity can affect the thermal performance of the wall structure to a certain extent.