We performed a series of collaborative experiments to clarify the effect of Mg++ on production of toxic-shock-syndrome toxin-1 (TSST-1) in various culture media. TSST-1 production was enhanced by adding ethylenediamine-tetraacetic acid (EDTA) at appropriate concentrations to brain-heart infusion and beef-heart medium. The magnitude of this effect depended both on the number of bacteria used to inoculate the media and on the sampling time. Large inocula prepared in media containing high levels of Mg++ introduced sufficient Mg++ to the growth medium to influence subsequent bacterial multiplication and toxin production. Small inocula of bacteria washed in Mg++-deficient medium before inoculation did not, however, multiply or produce toxin in Mg++-deficient medium. Maximal toxin expression occurred during late logarithmic phase, regardless of Mg++ concentration, and Mg++ appeared to control when late logarithmic stage of growth would be achieved. The toxin-enhancing effect of EDTA was reversed by adding excess Mg++ to treated medium.