Secondary caries is one of the main reasons for dental filling replacement. There is a need to obtain dental restorative material that is able to act against caries-inducing microorganisms. This study explores the antimicrobial properties of cetyltrimethylammonium bromide (CTAB) or dimethyldioctadecylammonium bromide (DODAB)-modified photo-cured experimental dental composites against Escherichia coli, Streptococcus mutans, and Candida albicans. The antimicrobial activity against Escherichia coli, Streptococcus mutans, and Candida albicans was assessed by using an Accuri C6 flow cytofluorimeter, and then analyzed using BD CSampler software (1.0.264). Bacterial/yeast surface colonization was carried out by using an GX71 inverted-optics fluorescence microscope equipped with a DP 73 digital camera. For bactericidal surface analysis of each sample type, simultaneous standardization was performed using a positive control (live cells) and a negative control (dead cells). A positive correlation between the increasing concentration of CTAB or DODAB and the dead cell ratio of Escherichia coli, Streptococcus mutans, and Candida albicans was revealed. In particular, CTAB at a 2.0 wt% concentration exhibits superior efficiency against pathogens (65.0% dead cells of Escherichia coli, 73.9% dead cells of Streptococcus mutans, and 23.9% dead cells of Candida albicans after 60 min). However, Candida albicans is more resistant to used salts than bacteria. A CTAB- or DODAB-modified experimental dental composite exhibits antimicrobial potential against Escherichia coli, Streptococcus mutans, and Candida albicans after 10 and 60 min of incubation, and the antimicrobial efficiency increases with the wt% of QAS in the tested material.