Stones of historical monuments exposed to the open air deteriorate over the course of time depending on physical, chemical, and biological factors acting in co-association. Among the biological factors, microorganisms play a key role in the deterioration process of stones. Detecting the level of microbial activity on stones is an essential step in diagnostic and monitoring studies of stone biodeterioration, and aids in controlling the performance of treatments applied to the stones. Therefore, this study aimed to develop a practical and rapid method for the determination of microbial activity on historical stones and use this method on the Mount Nemrut monuments (MNMs) (Adiyaman, Turkey). For that purpose, the fluorescein diacetate (FDA) hydrolysis method, frequently employed for soil environments, was adapted for the estimation and assessment of total microbial activity to understand whether microorganisms posed a potential risk for the biodeterioration of the limestones and sandstones of the MNMs. The traditional plate count method was also applied simultaneously to the same stone samples to compare and assist in the interpretation of the results of the FDA hydrolysis method, which relies on the quantitative determination of bacterial and fungal colonies in nutrient agar and malt extract agar medium, respectively. The results of the FDA hydrolysis and plate count methods showed consistency. The total microbial activity determined by the FDA hydrolysis method was low for both types of stone samples. In addition, the plate count method showed low bacterial and fungal counts on all of the samples. This revealed that microbial activity did not play an important role in the stone deterioration process on the MNMs, although different lichen species were frequently observed on both the sandstones and the limestones. Hence, further investigation must be undertaken for determination of their long-term behavior and effects on the stones of the MNMs. On the other hand, the results of the FDA hydrolysis and plate count methods showed correlation. Lower bacterial counts were observed when lower enzymatic activity was observed in the stone samples, and likewise, higher bacterial counts were observed when higher enzymatic activity was observed. Consequently, the application of the FDA hydrolysis method was determined to be reliable for the estimation of total microbial activity on historical stones. The method had obvious advantages in terms of its rapid measurement rate and sensitivity, even on small samples.