The authors have prepared monoclonal antibodies to each of the enteric adenoviruses types 40 and 41. Three different hybridoma cell lines were selected which produced antibody found to react by radioimmunoprecipitation with adenovirus (Ad) hexon antigens. One was specific for Ad40, another for Ad41, and a third one reacted with both types. When tested in an enzyme immunoassay against all 41 known human Ad types, the type-specific monoclonal antibody against Ad40 reacted homotypically, as did the monoclonal antibody against Ad41. In addition, these monoclonal antibodies neutralized the homologous enteric Ad type. The monoclonal antibody which reacted with both enteric Ad types also showed lower levels of reactivity with the Group C adenoviruses types 2,5, and 6. The monoclonal antibodies produced will provide a definitive means for rapid identification of specific Ad types, and will be useful in defining the relationship of enteric adenovirsuses to other types.