The monoclonal anti-idiotypic antibody (maId) 6G6.C4, directed against the carcinoembryonic antigen (CEA)-specific T84.66 immunoglobulin, was recently shown to act as a surrogate antigen for CEA in experimental animals. In this report, we have extended our studies. 1) The kinetics of complex formation in this CEA-specific idiotypic cascade were investigated using Biosensor technology. 2) Bacterial expression studies show that the mimicked epitope can be delimited to the A3 domain of CEA. 3) We cloned and characterized the genes coding for maId 6G6.C4. 4) Comparison of this epitope-bearing domain with the hypervariable region sequences of 6G6.C4 yields substantial amino acid similarity. Sequence homology between maId 6G6.C4 and anti-CEA antibodies binding to the T84.66 epitope led us to investigate the interaction of maId 6G6.C4 and CEA. Surprisingly, 6G6.C4 specifically binds to CEA but not CEA-related antigens in Western blots. 6G6.C4 and T84.66 recognize different epitopes on CEA. Our results suggest that the T84.66 epitope functionally mimicked by maId 6G6.C4 may be involved in the homophilic binding between CEA molecules, and that heterophilic interactions in the CEA-family are mediated by different binding sites. A model for the intermolecular adhesion of CEA is presented.