The aim of the present study was to establish an alternative methodology for testing the antibacterial effects of different amalgams. The vitality of mutans streptococci grown in vitro on various amalgam surfaces was monitored with a vital fluorescence staining technique using fluorescein diacetate and ethidium bromide. The in vivo effect of amalgam-non-gamma 2 fillings on the vitality of dental plaque was assessed with the same method and compared with samples originating from enamel. The median in vitro vitality of mutans streptococci was estimated as 70% on glass, 50% on Amalcapnon-gamma 2 and Sybraloy, 20% on Amalcap F and 10% on Neo-Silbrin. In vivo plaque vitality on enamel varied from 60 to 70%. In contrast, plaque sampled from non-gamma 2-amalgam surfaces revealed significant reductions in vitality with a minimum value of 25% of one day old supragingival plaque. The vital fluorescence technique was shown as an easy and quick method to assess the bactericidal effect against biofilm bacteria of dental materials in vitro as well as in vivo.