This MONIS 5 report focuses on freshwater crayfish and was funded by the Danish Environmental Protection Agency. The aim of the present study was to develop species-specific systems for tracking environmental DNA (eDNA) from nine species of freshwater crayfish in water samples. Among these nine species, only one (Astacus astacus) can be considered indigenous in Scandinavia, and among the other eight non-indigenous species; two (Astacus leptodactylus and Pacifastacus leniusculus) are frequently encountered in Scandinavian freshwater systems. The remaining seven non-indigenous species are rarely encountered in Scandinavia but have been recorded in the past from other places in Europe. It is our aim that the nine species-specific eDNA assays presented here will allow for continuous monitoring of both the one indigenous species, the two non-indigenous species more frequently encountered and help produce early warnings of the seven non-indigenous species that might disperse to Denmark.