At least 6 categories of pathogenic E. coli are undoubtedly recognized as being associated with enteric diseases. The importance of diarrheagenic E. coli is probably underestimated due to limited applications of available diagnostic methods in routine laboratories, especially in developing countries. Several strategies have been described for detection and characterization of diarrheagenic E. coli strains, mainly after the development of more advanced techniques. Thus, this chapter presents culture- and DNA-based methods, as well as immunological and biological assays to detect and characterize different diarrheagenic E. coli categories. Additional methods for confirmation and subtyping are also presented. The method of choice will depend very much on the facility of each laboratory, taking into account the sensitivity, specificity, reproducibility, reliability, cost, infrastructural resources and technical skills required by each method. Apart from diagnostic methods we also included some methods, which are more directed to research and epidemiological purposes, but may eventually be useful for a more specific characterization of the E. coli strains.