Some direct methods that can be used for the diagnosis of Lyme borreliosis are the culture, direct visualization or the detection of microbial DNA using polymerase chain reactions, in body tissues or fluids. Unfortunately, all these methods have a low sensitivity. There is a wide assortment of tests and antigens for indirect diagnosis, and the most recommended are ELISA tests and Western blot. The main inconvenient of these tests are the existence of shared serologic reactions, the variability of immune response and the difficult interpretation of results. Therefore, we propose the following guidelines for the diagnosis of Lyme borreliosis: For sero-epidemiological studies and to diagnose infection, antibodies should be determined in subjects with a compatible clinical picture, using an ELISA test that must be positive in at least two separate samples. All positive ELISA results should be confirmed with Western blot analysis, that must be interpreted using established criteria. Polymerase chain reactions should be used when they are available.