In Europe at least three human pathogenic species of Borrelia burgdorferi sensu lato are the causative agents of Lyme borreliosis. All three species have been isolated or detected by PCR from skin, CSF and synovial fluid of patients with skin lesions, neuroborreliosis and Lyme arthritis respectively. Studies using strains representing the three species as antigen for the immunoblot revealed that interpretation criteria depend strictly on the strain used as antigen. More than using certain species as antigen it is important to use strains (f.e. B. afzelii strain PKo) expressing certain immunodominant antigens like OspC and p17 which may not be expressed by other strains in vitro. Using strain PKo as antigen the two band criterium can be used without loss of too much sensitivity compared to using B. burgdorferi sensu stricto strain PKa2 and B. garinii strain PBi. The use of recombinant antigens allows selection of highly specific and combination of homologous antigens from different strains; however not all desirable antigens have been recombinantly expressed. Addition of p17 and p58 as antigens may improve the sensitivity of the hitherto described recombinant antigen immunoblots containing the antigens p83/100, p39, OspC and the p41 internal fragment.