The cell envelope of Helicobacter pylori contains lipopolysaccharide (LPS), the O-chain of which expresses type 2 Lex and Ley blood group antigens, which mimic human gastric mucosal cell-surface glycoconjugates and may contribute to the survival of H. pylori in gastric mucosa. Here we describe the generation of monoclonal antibodies specific for Lex and Ley blood group determinants and the characterization of their binding properties using purified, structurally defined H. pylori LPS, synthetic glycoconjugates, and H. pylori cells. Analysis of oligosaccharide binding by SPR provided a rapid and reliable means for characterization of antibody affinities. One of the antibodies, anti-Lex, was of IgG3 subclass and had superior binding characteristics as compared with the commercially available anti-Lex IgM. These antibodies could have potential in the immunodiagnosis of certain types of cancer, in serotyping of H. pylori isolates, and in structure-function studies.