HCV transmission is commonly derived from blood transfusions. Some different aspects, such as location, infection prevalence, and genotype distribution, may affect the occurrence of HCV in blood donors. The blood donors have already been screened regularly for their anti-HCV serology, yet the test for HCV RNA has not been done yet. In this study, we aim to investigate the manifestation of HCV in Tuban by detecting HCV RNA from sera negative for HCV antibodies in blood donors. The blood donors from Tuban Red Cross Indonesia were recruited for a questionnaire interview and testing for HCV antibodies and HCV nucleic acids. Anti-HCV was serologically detected using ELISA. Nested PCR was used to amplify HCV-RNA in the NS5B and 5’UTR regions. The genotype or subtype of HCV is determined by direct sequencing followed by phylogenetic analysis. A total of 100 blood samples were collected. The HCV RNA positive rate was 6% in sera-negative anti-HCV blood samples. Furthermore, the genotyping resulted in 4 samples being dominantly HCV subtype 1c (66,67%); the other 2 samples were subtype 2a and type 1 (each counted as 1 individual, 16.67%, respectively). The serological test for HCV antibodies has been shown to be less sensitive than the nucleic acid amplification test. The detection of genotype 1c as a major HCV genotype circulating in the Tuban area may help to anticipate HCV transmissions and facilitate better medical treatment with respect to HCV carriers.