Staphylococcus aureus is a food-borne pathogen well-known by its ability to adhere and form biofilms in abiotic surfaces. One of concern about this bacterium is the growing reports that describe its isolation from food-contact surfaces in Food Services Units. S. aureus biofilms act as continuous sources of contamination and enhances its persistence in environments of food processing. The aim of this study was evaluate the capability to form biofilm in strains isolated from food-contact surfaces of food processing environments of hospitals by different detection methods. The biofilm formation was assessed in 10 non-clones S. aureus coagulase positive selected from 60 previously isolates using microtiter plate method (MtP), Congo red agar (CRA) icaA and icaD gene detection by PCR. The strains S. epidermidis ATTCC 12228 and S. aureus ATTCC 25923 were used as negative and positive (strong producer) control for biofilm formation, respectively. According MtP results, considering the optical density (OD) measurement of the test and negative control isolate (ΔOD570 nm), the ten strains tested were classified as strong biofilm-producers, (4x ODcontrol< OD). All strains showed the amplified fragments of 188bp and 198bp, corresponding to icaA and icaD genes in PCR assays, respectively. Among the strains, seven formed rough-black colonies on CRA plates and were considered to be biofilm producing strains, while three formed red colonies and were evaluated as moderate biofilm producing strains. These findings show that the molecular detection of icaA and icaD genes does not the phenotypic expression and suggest that environment factors might play a major role in triggering biofilm production.