The adaptive reuse of historic buildings, as well as the maintenance of their optimal indoor environments for structural and collections preservation, are inherently complex, multifaceted goals. Moreover, indoor environments do not only affect collections and building—the occupants’ satisfaction must also be considered. In this study, a field investigation of Indoor Environmental Quality (IEQ) particularly focused on thermal comfort following a Post-Occupancy Evaluation (POE) process applied to a case study where a combination of microclimate monitoring and questionnaire surveys were carried out during the problematic cooling season in the hot and humid climate of San Antonio, Texas (USA). Mission Concepción de Acuña (part of a UNESCO World Heritage Site) is a unique historic settlement that includes a limestone church which reflects a similar typology to multiple religious buildings distributed across the southwestern USA. Results of these investigations show that: i) the current indoor environment did not always achieve occupants’ thermal satisfaction, displaying thermal discomfort when the indoor/outdoor thermal leap is larger (August and September); ii) Thermal Sensation Vote (TSV) Predicted Mean Vote (PMV) values were considerably different throughout the monitoring period, with TSV mostly included within the (−1, 0, +1) band and PMV mainly within the (−2, −3) band; iii) impact of occupant age, gender and clothing on TSV values is limited. Finally, outcomes of this article underline the limitations of Fanger’s model when applied to such typology of buildings, highlighting the necessity for more investigation in this topic.