In efforts to mitigate greenhouse gas emissions in the construction sector, policies promoting energy efficiency improvements in existing buildings are being implemented. Accurately assessing the impact of energy retrofit strategies requires precise determination of thermal envelope performance through energy audits. Aerial thermography, employing unmanned aerial vehicles (UAVs) equipped with infrared thermal cameras (IRT), is an underused method overcoming limitations in energy audits of large buildings or complexes. This approach offers an elevated perspective, providing an overview of the building’s thermal patterns, aiding in the identification of thermal envelope defects. By measuring temperature distribution, even in inaccessible envelope areas, the severity of identified defects can be estimated. Beyond detecting thermal bridges, air leaks, and moisture, aerial thermography is valid for estimating thermal transmittance (U-value) of enclosures. The aim of this research is to evaluate this method to calculate the U-value of the façade of an university building located in a Subtropical Mediterranean climate, during a typical full day in different seasons of the year. Concurrently with aerial thermography, the thermometric method (THM) was applied to compare results and refine the operational conditions during U-value measurement using different statistical indices and uncertainty analysis. The results demonstrated the validation of airborne thermography as a method for U-value calculation under specific meteorological and operational conditions, overcoming the common physical limitations of traditional building audit methods.