Environmental DNA (eDNA) offers an opportunity for detection and monitoring of invasive vertebrate species. This approach is ideal for species that are semi-aquatic or terrestrial species that frequently interact with water. For example, feral swine (Sus scrofa), which are invasive across a significant portion of the U.S., are not semi-aquatic but they routinely wallow in water. Burmese pythons (Python bivittatus), which are invasive in south Florida, are a semi-aquatic species. Both species cause significant impacts to native species and ecosystems and thus are of interest to managers. We designed species specific primers, tested various approaches to DNA isolation and concentration, and quantified our limits of detection and quantification. We also assessed persistence of eDNA in the target environment. Our work demonstrates the importance of pen tests with target species to optimize the eDNA assay and further field testing as a pilot study before full studies may be successful. In our pilot field studies we implemented modeling to assess what factors influenced our probability of detection and the laboratory methods that most influenced detection as well. Finally, we developed a method for rapid and easy field collection of samples. The field of eDNA is rapidly developing and much of the knowledge is housed within laboratory personnel rather than in publications. It is my goal to have an open discussion about laboratory and field techniques to further the sharing and integration of knowledge between practitioners and those interested in applying these methods in wildlife conservation.