Bactericidal efficacy of seven different disinfectants (chlorhexidine gluconate, povidone-iodine, glutaraldehyde, benzalkonium chloride, sodium hypochloride, ethoxydiamino acridine, the mixture of "12% ethandiol + 0.5% pentanediol + 7.5% didecyldimethylammonium cloride") commonly used at Osmangazi University, Medical Faculty Hospital, was investigated against frequent nosocomial bacterial pathogens (19 Gram-negative and 25 Gram-positive strains) by using modified Kelsey-Sykes method. Chlorhexidine gluconate, povidone-iodine and glutaraldehyde were determined as the most effective disinfectants against all of the pathogens. There was no significant difference in terms of biocide resistance between antibiotic sensitive and resistant strains. All of the P. aeruginosa strains were found to be resistant to 0.01% benzalkonium chloride and 0.25%, 0.5% and 1% concentrations of "12% ethandiol + 0.5% pentanediol + 7.5% didecyldimethyl ammonium cloride" mixture. One vancomycin resistant E. faecium strain and one extended spectrum beta-lactamase (ESBL) positive K.pneumoniae strain were found to be resistant to 0.55% sodium hypochloride. Ethoxydiamino acridine showed little or no activity against all tested strains.