Abstract The effect of metallothionein (MT) on lymphocytes was studied in vitro. Rabbit MT induced the proliferative responses of mouse splenocytes at concentrations of 1–25 μg/ml. MT synergistically enhanced Con A- or LPS-induced proliferative response of splenocytes. A similar effect was also observed for rabbit splenocytes at a similar concentration. Free heavy metals such as Cd and Zn only weakly stimulated splenocytes. 2-mercaptoethanol (2-Me) abrogated the effect of MT, suggesting that thiols in MT play an important role for splenocyte response. MT stimulated splenocytes from MT-knockout mice as well as those from normal control mouse. The responder cells for MT stimulation were B cells and MT also induced B cell differentiation to produce Ig. MT induced the calcium influx of B cells, but not T cells. These results indicate that MT has a potent immunomodulating activity, especially on B cells.