Volume 116, Issue 3, Pages 636-642 (March 1999) Real-time detection system for quantification of hepatitis C virus genome  Tomoko Takeuchi, Asao Katsume, Takeshi Tanaka, Aki Abe, Kazuaki Inoue, Kyoko Tsukiyama-Kohara, Ryuji Kawaguchi, Satoshi Tanaka, Michinori Kohara  Gastroenterology  Volume 116, Issue 3, Pages 636-642 (March 1999) DOI: 10.1016/S0016-5085(99)70185-X Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 1 Schematic structure of DNA templates used to synthesize standard HCV RNA. Vectors were constructed as described previously.15 PKI5 codes sequences from HCV genotype 1b. PKII5 codes sequences from HCV genotype 2b. Gastroenterology 1999 116, 636-642DOI: (10.1016/S0016-5085(99)70185-X) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 2 Serum ALT levels and HCV-RNA quantity in the 4 patients treated with IFN-α. (A) CR-SR case; and (B–D) CR-Rel cases. ●, ALT levels; ○, RTD-PCR (set 2 primers and probe) titers; □, Amplicor monitor titers. Gastroenterology 1999 116, 636-642DOI: (10.1016/S0016-5085(99)70185-X) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 3 Amplification plots of synthetic HCV RNA. Tenfold serial dilutions of target HCV-RNA were prepared in triplicate, reverse transcribed and amplified using TaqMan probe, and detected using ABI Prism 7700 sequence detector. For each dilution, Δ normalized reporter (Rn) is plotted against each cycle number. (A) Fluorescence units using set 2 primers, probe, and genotype 1b standard HCV RNA. (D) Fluorescence units using set 2 primers, probe, and genotype 2b standard HCV-RNA. B (genotype 1b) and E (genotype 2b) show Ct values19,20 plotted against relative copy number for two genotypes of HCV. C (genotype 1b; set 1) and F (genotype 1b; set 2) show double-strand DNA products of PCR reaction (53 cycles) with ethidium bromide. The calculated Ct value is predictive of numbers of copies of target RNA present in sample. bp, base pairs. Gastroenterology 1999 116, 636-642DOI: (10.1016/S0016-5085(99)70185-X) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 4 Correlation between serum HCV-RNA levels determined using RTD-PCR and those determined using (A) bDNA assay and (B) Amplicor monitor using set 1 primers and probe. The vertical dotted line indicates the detection limit of (A) bDNA assay and (B) Amplicor monitor. Gastroenterology 1999 116, 636-642DOI: (10.1016/S0016-5085(99)70185-X) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 5 Correlation between serum HCV-RNA levels determined using RTD-PCR (set 2 primers and probe) and those determined using Amplicor monitor. (A) Genotype 1b; (B) genotype 2a; and (C) genotype 2b. Gastroenterology 1999 116, 636-642DOI: (10.1016/S0016-5085(99)70185-X) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 6 Prevalence rate of HCV RNA in HCV-positive or -negative control sera. ASC, seropositive for HCV antibody and normal ALT levels for more than 6 months; CH, patients with chronic HCV; LC, patients with HCV liver cirrhosis; HCC, patients with HCV hepatocellular carcinoma; HBV carrier, patients with chronic HBV; Non-B, Non-C, HBV and HCV marker–negative patients with hepatitis; and Healthy, HBV and HCV marker–negative and healthy donor. Gastroenterology 1999 116, 636-642DOI: (10.1016/S0016-5085(99)70185-X) Copyright © 1999 American Gastroenterological Association Terms and Conditions