Background: Considering the increased antimicrobial resistance to Staphylococcus aureus and also the importance of vancomycin as a treatment, the aim of this work was to determine the minimum inhibitory concentration (MIC) of vancomycin, the pattern of antibiotic susceptibility, removing the isolates producing beta-lactamase enzyme and determining the risk factors for prevalence of methicillin-resistant Staphylococcus aureus (MRSA) isolated from clinical samples in Shahid Beheshti Hospital of Kashan-Iran.Materials and Methods: This descriptive study was carried out on 150 Staphylococcus aureus isolates. The sample strains were identified using gram staining, catalase test, tube coagulase test, growth on manitol salt agar medium and the DNAase test. The antibiotic sensitivity test was determined by disk diffusion method on nine antibiotics. MIC of vancomycin was determined using Broth Microdilution Method. The production of beta-lactamase enzyme was performed using acidometry method. Results: Among the 150 Staphylococcus aureus isolates, MIC value was 0.5-4 mg/mL. The highest MIC of vancomycin was 2 mg/mL. All of isolates were sensitive to vancomycin. Mean age of patients was 31.3+25.8. Penicillin and vancomycin revealed the highest and the lowest resistant rate, respectively (93% and 0%). Resistance to methicillin was 62%. Eighty-seven percent of the isolates produced enzyme beta lactamase. Risk factors for prevalence of methicillin resistant Staphylococcus aureus were high age and previous antibiotic therapy. Conclusion: Considerining the vancomycin as the selective antibiotic, on following the preventive guideline, preparing the antibiogram and adding MIC test are recommended.