Two European species of the genus Eteobalea Hodges are being tested as classical biological control agents for toadflax (Linaria spp.) in Canada. Permits for the release of these species apply only to an Italian population of E. serratella Treitschke and a Serbian population of E. intermediella Riedl. When these species are imported as larvae or pupae from an area where they are sympatric, such as around Rome, they must later be separated in the laboratory. This step is essential to establish a pure laboratory colony of the correct species, both to increase the rate of breeding success and to comply with conditions of the importation permit. Although the male and female genitalia and the egg chorion have good diagnostic characters, these characters are not useful for identifying immature stages or newly emerged adults before mating and oviposition. This problem has hampered the establishment of rearing colonies of these two species in Canada. We describe the development and application of four polymerase chain reaction – restriction fragment length polymorphism assays that have been used to distinguish between these two species. If and when either of these species becomes established in the field, the tests presented here should also prove useful for monitoring populations.