Abstract:A high-throughput polymerase chain reaction (PCR)-based enzyme-linked oligonucleotide-sorbent assay ELOSA)was developed for use in the diagnostic testing of serum from patients who may be infected with different hepatitis C virus (HCV) genotypes. Twelve genotype-specific 5 -aminated DNA-coated probes were designed based on the variable 5 -untranslated region sequences of the HCV genotypes 1–6. Using 100 clinical serum samples, the performance of the PCR-ELOSAmethodwascompared with Roche’s COBAS Amplicor HCV Monitor V2.0 assay and the VERSANT HCV genotype assay (LiPA), and the overall agreement was 99% at the level of HCV genotypes with a detection range of 2.0×102 to 1.0×107 IU/ml for PCR-ELOSA. The PCR-ELOSA was more comprehensive as demonstrated by the fact that ?20% of the samples with different subtypes could be discriminated by this method but not by LiPA. In addition, the PCR-ELOSA system showed high accuracy (CV?6.36%) and even higher reproducibility (CV?5.55%). Thus, this novel PCR-ELOSA system provides a sensitive and versatile alternative to current HCV detection assays.