BACKGROUND: The detection of antibody to hepatitis C virus (anti-HCV) is the most useful method to investigate past or current HCV infections. We performed a clinical evaluation of ADVIA Centaur HCV assay, a new third generation assay for the qualitative detection of IgG antibody. METHODS: Included in the study were a total of 323 samples (108 positive and 215 negative), for which HCV reverse transcription polymerase chain reaction (RT-PCR) was requested. ADVIA Centaur HCV assay (Bayer Healthcare LLC, Diagnostics Division, Tarrytown, NY, USA) was compared with a currently available and widely used AxSYM HCV version 3.0 assay (Abbott Laboratories, Abbott Park, IL, USA). Samples with discrepant results were retested with each assay, and further tested with a recombinant immunoblot assay (RIBA, LG HCD Confirm, LG Chemical Co., Seoul, Korea). Reproducibility of Centaur assay was evaluated in five groups (two samples in each group) with different index values. RESULTS: The overall concordance rate was 91.6% (296/323) between Centaur and AxSYM assays. It was 100% (108/108) in RT-PCR positive samples and 87.4% (188/215) in RT-PCR negative samples. Discrepant samples (8.4%, 27/323) were all RT-PCR negative, and all except two were Centaur negative and AxSYM positive. In discrepant samples, RIBA showed negative results except for two samples with indeterminate results. The sensitivity and specificity of Cenyaur assay were 98.1% and 65.6%, and the respective figures for AxSYM assay were 98.1% and 54.9%. Reproducibility of Centaur assay was satisfactory. CONCLUSIONS: The overall concordance between Centaur and AxSYM assays was satisfactory. Sensitivity and specificity of Centaur assay were equivalent to or better than those of AxSYM assay. ADVIA Centaur HCV assay seems to be a reliable and useful method for the detection of anti-HCV in clinical laboratories.