Using a mice in a tetanus vaccination system, we have examined the mode of action of the immunological adjuvant, aluminium hydroxide (Al(OH)3), on antibody titers as measured by an ELISA-method. A profound effect of the adjuvant was observed after primary immunizations, increasing titers about 100-fold compared to plain (not adjuvanted) controls. However, no effect on titers could be attributed to a content of Al(OH)3 in the vaccines when used for booster immunizations, as the increase in titers after boosting was either the same for groups given plain or adsorbed toxoids, or even higher in the groups receiving the plain vaccine. Thus, the effect of Al(OH)3 as an adjuvant seems to be exerted mainly on the primary antibody response. Investigating the mode of action by the adjuvant, we adsorbed a certain amount of toxoid onto varying doses of Al(OH)3. An antigen ELISA revealed that all toxoid had been adsorbed to the Al-gel in the preparations. The antibody titers after immunization showed a significantly higher response to toxoid adsorbed to the higher amount of adjuvant, indicating that this effect was most probably due to free, not antigen-bound, Al-particles. Moreover, this effect could be inhibited when BSA was added in excess to the preparation, thereby blocking the free residues on the Al-gel.