In this study the lipids of Borrelia burgdorferi, the causative agent of Lyme disease, were analyzed. Lipids comprise about 25-30% of the cell dry weight. The lipid fraction could be separated by HPTLC into 11 components. Staining of these components revealed two glycolipids and two phospholipids. The glycolipids represented about 50% of the total lipids and comprised only galactose as monosaccharide constituents. By means of mass spectrometric and gas chromatographic analysis both glycolipids could be identified as alpha-galactosyl-diacylglycerolipids with different fatty acid compositions. The phospholipids were identified as phosphatidylcholine and phosphatidylglycerol. Immunoassays with sera from patients with Lyme disease showed antibody reactivity only to the glycolipids, which was present in all stages of the disease. Other lipid components seemed to be non-immunogenic in Lyme disease. The glycolipids of B. burgdorferi may be, thus, considered promising candidates for diagnosis and possibly also for vaccination.