The aim of our study was twofold: to immobilize an organosilicon quaternary ammonium salt (3-(trimethoxysilyl)-propyldimethyl-octadecyl ammonium chloride, Si-QAC) on the surface of pure titanium and to investigate the antimicrobial activity of Si-QAC-immobilized titanium against microbial adherence and biofilm formation. The results of ToF-SIMS analysis of Si-QAC-titanium suggested the possibility of immobilizing Si-QAC on titanium surface through Ti-O-Si coupling, and that Si-QAC treatment significantly reduced both the adherence and colonization of Candida albicans and Streptococcus mutans isolates. The antimicrobial activity was achieved through at least two mechanisms: the first was attributed to the octadecyl alkyl chain which inhibited initial adherence, and the second was attributed to the quaternary ammonium salt which killed initial adherent cells as well as retarded or inhibited subsequent microbial growth. Further, thermocycling did not significantly reduce the antimicrobial activity of Si-QAC-titanium, and no significant cytotoxicity of Si-QAC-titanium was observed in either cell viability test or proinflammatory cytokine production test using human gingival fibroblasts. These results, taken together, favorably suggested that Si-QAC treatment would be a helpful means to inhibit dental plaque or denture plaque formation.