Lyme borreliosis (LB) is the most prevalent tick-borne zoonosis in Europe and North America. Borrelia burgdorferi, the causative agent of LB is a gram-negative, microaerophilic bacterium which belongs to the Family Spirochaetaceae. Infection with B. burgdorferi can result in dermatological, neurological, cardiac, and musculoskeletal disorders. The diagnosis of LB is ascertained by clinical findings and microbiological tests. Microbiological tests involve direct and indirect methods. In direct methods, detection of either the microorganism by direct microscopic examination and culture or its nucleic acid by PCR is intended. As for the indirect methods, they are used in order to detect the antigenic compounds of microorganism and specific antibodies in patient's serum by serological tests such as ELISA, IFA, and Western Blot. ELISA is the most frequently used diagnostic test in LB. However, the diagnosis of the LB should be done by evaluation of history and clinical findings of the patient as well as laboratory findings together. Although the serologic tests are not mandatory so much for diagnosis of the disease at the first stage, laboratory findings are needed in addition to the clinical findings at the second and third stage. The tests performed without clinical findings may cause confusion. In the presence of clinical findings, the clinical picture should be verified with determination of the stage of the infection, organ system affected and appropriate diagnostic tests.