A recently described PCR test for the identification of Neospora caninum and Toxoplasma gondii has been further developed and optimized in view of its practicability for routine diagnostic application. The N. caninum-specific PCR was adapted to the diagnostic operating standard of the T. gondii-specific PCR in that the uracil DNA glycosidase system was introduced, which eliminates potential carry-over contaminations of amplified target DNA from previous reactions. Furthermore, both PCR tests were optimized by including a DNA hybridization immunoassay based on the use of the commercially available Gen-eti-k DEIA kit. This assay allowed highly sensitive and specific detection of respective DNA amplification products and thus substantially facilitated the reading and interpretation of the test results.