Background. Coagulase-negative staphylococci (CoNS) have emerged as an important opportunistic pathogen isolated in hospital. Several species of CoNS have been implicated in human infections and disease especially in patients with poor health status. Methods. A total of 71 clinical strains of CoNS were isolated from dialysis fluid and needles in a dialysis unit and characterized. Susceptibility to antibiotics, biofilm production and molecular typing by pulsed-field gel electrophoresis (PFGE) were achieved. Results. The main isolated CoNS strains were Staphylococcus epidermidis (45%), Staphylococcus hominis (14%) and Staphylococcus haemolyticus (12.7%). The susceptibility profile of all strains revealed high resistance level to penicillin and oxacillin. PCR detection of oxacillin resistance gene (mecA gene) revealed a higher percentage of positive strains than the classic test (ATB Staph). Slime production test was positive in 60.6% of CoNS strains. PFGE analysis showed the presence of 69 restriction profiles clustered in 56 patterns. Conclusions. Profiles of all isolates were generally heterogeneous, suggesting independent circulation with some evidence of cross-transmission. (C) 2006 IMSS. Published by Elsevier Inc.