BACKGROUND Hepatitis C virus (HCV) is a leading cause of chronic hepatitis in dialysis patients. The diagnosis of HCV infection in these patients is predominantly based on laboratory tests because of the specificity of the clinical course of the disease. AIM The present prospective study aimed at determining very accurately the prevalence rate of HCV infection in patients on dialysis by simultaneously testing them for anti-HCV and for HCV RNA levels. MATERIALS AND METHODS For the present cross-sectional longitudinal study we recruited and followed up 93 patients from St George University Hospital Hemodialysis Unit between July 2013 and December 2014. All patients were tested for anti-HCV and HCV RNA. The anti-HCV negative patients were tested for anti-HCV and HCV RNA at least twice at intervals of 6 months or more (up to 12 months). Anti-HCV antibodies were identified using a third generation ELISA assay. Commercial kits for real-time polymerase chain reaction (RT-PCR) were used to detect HCV RNA in the plasma and mononuclear cells. Aminotransferase and gammaglutamyl transpeptidase levels were studied to find if liver inflammation was present. RESULTS The total seroprevalence in 68 patients was 20.6% (14). Of these, 10 patients were viremic (HCV RNA+/anti-HCV+), and 4 patients (5.9%) had discordant results (anti-HCV+/HCV RNA-). Acute hepatitis was detected in one patient. Duration of dialysis in HCV viremic patients was longer than that in aviremic patients (p=0.005). CONCLUSIONS The present study suggests that HCV infection in dialysis patients can be diagnosed more accurately if these patients are tested using two diagnostic methods - a serological test and a biomolecular assay. Further studies with larger sample size may prove the feasibility of such approach for all dialysis patients in this country.