Freshly recovered pulmonary alveolar macrophages (AM) and the AM-derived established cell line, MH-S, have previously been shown to be highly suppressive of in vitro IgM anti-sheep erythrocyte (SRBC) responses. Supernatants obtained from cultures of AM incubated with antigen-stimulated lymphocytes or from the MH-S cell line alone have also been shown to be suppressive when added to the in vitro antibody-forming system. In order to determine if AM and MH-S cells, owing to their mucosal location, could differentially regulate antibody responses including immunoglobulin isotypes other than IgM, an in vitro system for the detection of cells producing IgG, IgA and IgE anti-2,4 dinitrophenol (DNP) antibody was developed. These studies demonstrate that AM, MH-S cells, and MH-S culture supernatants suppress the in vitro generation of IgM, IgG, IgA and IgE anti-DNP spot-forming cells (SFC). No apparent differential regulation of any of the four murine IgG anti-DNP antibody subclasses was observed. Time-course experiments suggested that optimal AM- and MH-S-mediated suppression occurred 18 hr after culture initiation. Both AM and MH-S cells suppressed IgM and IgG anti-DNP antibody responses in a dose-related manner, suggesting that MH-S is a good model for the study of AM-mediated immunoregulation.