Abstract MBr1 is a murine monoclonal antibody, defining a saccharidic epitope [CaMBr1] of a human tissue-specific, tumor-associated globoside, present on the mammary carcinoma cell line MCF-7. The same epitope is shared by glycoproteins present on normal and neoplastic mammary epithelial cells, and by mucins from some ovarian cyst fluids. We have used MBr1 as the monoclonal antitumor antibody in an idiotypic sequence of immunizations in order to obtain and characterize "internal images" of the original epitope to be used as substitutes of the nominal antigen in serologic immunoassays. Two monoclonal anti-idiotypic antibodies (beta-1 and beta-2), which reacted with paratope-related idiotopes on MBr1, were obtained. The analysis of the antigenic and immunogenic properties of these molecules by both "antigen" and "antibody" competition assays provided evidence that both beta-1 and beta-2 bear "internal images" of the MBr1-defined epitope. Moreover, when injected in mice and rabbits both beta-1 and beta-2 induced anti anti-idiotypic antibodies, which mimicked MBr1 in binding MCF-7 as well as normal and neoplastic mammary gland epithelial cells. These data are discussed in terms of their possible application to the production of tumor-associated antigen substitutes and their use in serologic immunoassays.