Polichloride vinyl (PCV) is a widely used thermoplastic polymer, also in the production of medical devices. In the present study we assess the influence of EDTA in vitro on the biofilm structure formed by Staphylococcus epidermidis isolates on PCV biomaterials (Nelaton and Thorax catheters). The 6 strains of S. epidermidis were isolated from nasopharynx of hospitalised patients. It was found that all isolates were able to form the biofilm on both PCV biomaterials, irrespective of adhesion properties (cell surface properties, ability to slime production, minimal time needed for adhesion). The EDTA showed bacteriostatic effect against planktonic cells of the isolates (MIC = 0.25–0.5 mmol/l; MBC = 10.0–>25.0 mmol/l; MBC/MIC = 20, 30, 40, >50). The adhesion process and also formation of the biofilm was inhibited by EDTA at concentrations 1.0–2.0 mmol/l (2–8 × MIC). The eradication of the mature biofilm was achieved at 2.0–4.0 mmol/l EDTA (4–8 × MIC) for two strains, while for the other four isolates, concentration of EDTA needed for eradication effect was >32 mmol/l (> 128 × MIC). Data obtained in this paper suggest that EDTA may be regarded as a useful agent preventing formation of the S. epidermidis biofilm on PCV biomaterials.