Quantitative real-time PCR (qPCR) analysis of environmental DNA (eDNA) has become a sensitive and cost-effective tool for monitoring freshwater ecosystems and for the early detection of invasive fish species. The European catfish (Silurus glanis L.) is one of the largest apex freshwater predator worldwide and can exert a significant negative impact on local fauna in non-native areas, therefore its early detection is crucial for effective management. In this study, qPCR and standardized sampling methods (i.e., multi-mesh gill nets, both benthic and pelagic, and point-abundance electrofishing) were compared to assess the presence of S. glanis in 15 lakes in northern Italy. The species was absent in eight lakes, whereas it was detected in seven lakes using standardized sampling methods. Among these, eDNA analysis detected the species in only one lake, suggesting limited persistence of its DNA in the environment. To further investigate this aspect, we experimentally evaluated the release and degradation dynamics of S. glanis eDNA in indoor tanks. Our results show that eDNA concentration declined rapidly after fish removal, and S. glanis became undetectable by qPCR within six days. Our results suggest that, although eDNA analysis is a promising approach, it cannot yet replace standardized sampling methods for detecting and monitoring elusive fish species. Furthermore, understanding the release and degradation dynamics of genetic materials in the environment is crucial for the correct interpretation of eDNA results.