There is an essential requirement for highly sensitive tools that will differentiate nematode parasites of animals and plants to the species level. For studying host range, genetic variation, virulence and resistance, the availability of well defined populations is vital. Many nematode species cannot be identified with certainty using traditional morphological or morphometric techniques. This is particularly the case for the more accessible developmental stages that, depending on the particular group concerned, live as eggs and larvae in the environment or as micro-filariae that circulate in the blood or inhabit the skin. Morphological identification of these stages requires specialized expertise and is extremely time consuming. Immunological assays have their place in nematode identification but they do not discriminate between current and previous infections, an essential requirement in many epidemiological and prevalence studies. In addition to being highly sensitive, DNA-based methods of detection define present over past infection and are not dependent on the parasite stage. Many types of methodology are available for the detection and definition of nematode DNA. This paper reviews these methods citing examples that have been used with success in the laboratory as well as the field.