The insulin-degrading enzyme (IDE), a Zn2+-dependent metalloprotease, degrades several physiologically important peptides including A, a peptide involved in the pathogenesis of Alzheimers disease. Enhancing IDE activity may be therapeutically beneficial because it would lead to increased clearance of A thereby minimizing its toxic effects. In this study, small-molecule activators of IDE were identified by means of a high-throughput screen, using rat IDE and a synthetic fluorogenic reporter. The results demonstrate that IDE activators exhibit substrate and species specificity, which was observed upon comparing the effects of activators on various IDE orthologs and in the context of distinct reporters. The results of a high-throughput screen aimed at identifying small-molecule activators of human IDE using a novel A synthetic fluorogenic reporter are also discussed. This study provides insight into the various considerations that should be taken into account during the design of high-throughput screens aimed at identifying IDE activators. This study also indicates that activators can be substrate-specific thereby minimizing the impact on other IDE substrates including insulin.