Nosocomial infections account for one of the most serious complications in hospitalized patients around the world. Surgical site infections have significant economic implications, and surgical antisepsis plays an important role in such processes.With prior approval by the Institutional Review Board and informed consent, 10 volunteers were randomly assigned to 3 protocols on hand antisepsis: protocol A (chloroxylenol 3%), protocol B (benzalkonium chloride at 1%), and protocol C (ethyl alcohol 61%, 1% chlorhexidine gluconate). Smears from both hands were cultured after each hand pro tocol (t0) and at the end of suturing (t1). Colony forming units were counted (CFUs on blood agar dishes) with digital counting software (Open CFU). Friedman test was used to compare the mean values among the groups, and a Bonferroni correction was made to determine the dissimilar group, with a P = 0.015.At t0 for protocol A the CFU count was 82.8 ± 1.3; protocol B was 9.7 ± 30; protocol C was 0.1 ± 0.3 (P < 0.001). At t1 for protocol A the CFU was 80.7 ± 89.4; protocol B was 7.5 ± 32; protocol C was 0.0 ± 0.0 (P < 0.001). No adverse events were present among the subjects.Ethyl alcohol at 61% with 1% chlorhexidine gluconate showed higher efficacy than the traditional washing antiseptics.