Protein antigens elicit humoral responses in mice that consist predominantly of IgG1 antibodies. We have now investigated the ability of IL-12, a cytoklne reported to augment IgG2a antl-hapten responses through activation of T_h1 cells, to alter antibody responses to hen eggwhite lysozyme (HEL). The normal response of BALB/c mice to HEL is highly restricted to lgG1 expression and therefore provides an excellent system for determining effects of cytoklnes on expression of other isotypes. Seven days after immunization, IL-12-treated mice demonstrated greatly elevated HEL-speciflc IgG2a antibody levels and suppressed IgG1 production, while PBS-treated control mice showed a typical lgG1-restricted response. On day 28, IL-12-treated mice showed heightened serum antibody levels of both isotypes. Delaying cytoklne treatment until after the typical IgG1 anti-HEL response had already been established also led to significant elevation of serum IgG2a antibody levels. These effects correlated with increased IFN-γ production; however, administration of IL-12 plus anti-IFN-γ had little influence on lgG2a enhancement, although it did relieve the early lgG1 suppression. Furthermore, the differential effects of IL-12 on Isotype expression did not correlate with time; in fact, IgG2a enhancement correlated with loss of IgG1 suppression. Our findings indicate that (I) IL-12 reproduclbly Induces large amounts of IgG2a HEL-speclflc antibodies In vivo; (II) it can alter isotype profiles of both primary and secondary responses; and (III) its effects on humoral immunity are not completely explained by Induction of T_h1 cell-derived IFN-γ.