Dynamic Range and Reproducibility of Hepatitis B Virus (HBV) DNA Detection and Quantification by Cobas Taqman HBV, a Real-Time Semiautomated Assay

Lindh, Magnus; Hannoun, Charles

doi:10.1128/jcm.43.8.4251-4254.2005

Cited by 25 publications

(20 citation statements)

References 25 publications

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“…The evaluation of quantitation differences in clinical samples demonstrated that CAP-CTM has an excellent correlation with the endpoint PCR system CAHBM, a finding in agreement with other studies (12,13,19,35). Differences for averaged log 10 values between CAP-CTM and CAHBM were below Ϯ0.5 IU/ml and even less when only samples within the endpoint PCR dynamic range are considered.…”

Section: Discussionsupporting

confidence: 75%

“…Reliable PCR results also depend on sample preparation. Nucleic acid extraction from biologic specimens is technically demanding and a potential source of run-to-run variability and sample contamination (12,19,35). As a result, there is a growing need for automated sample processing systems to provide accurate quantification of nucleic acid for clinical purposes.…”

Section: Discussionmentioning

confidence: 99%

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COBAS AmpliPrep-COBAS TaqMan Hepatitis B Virus (HBV) Test: a Novel Automated Real-Time PCR Assay for Quantification of HBV DNA in Plasma

et al. 2007

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Hepatitis B virus (HBV) infection is a major cause of chronic liver disease. It is estimated that nearly 2 billion people are infected worldwide by HBV and that more than 350 million have persistent and chronic infection (38). HBV carriers have a high risk of developing long-term sequelae of hepatitis B, including cirrhosis and hepatocellular carcinoma that, in countries such as Italy with a moderate prevalence of HBV infection, account for nearly 25% of the indications for liver transplantation in reference centers (29). Recent advances in antiviral therapy, based on the development of new and more powerful nucleos(t)ide analogues, have dramatically improved chronic hepatitis B management, including the prevention of allograft reinfection in those patients undergoing liver transplantation for HBV-related disease.The success of antiviral therapy has been supported by the introduction of highly sensitive tests for monitoring HBV DNA. Molecular tests help to determine the activity of HBV infection, the selection of patients for treatment, and the efficacy of antiviral therapy, identifying the development of HBV drug-resistant strains (16,36,38).Several assays based on PCR are currently commercially available for HBV DNA. The recently introduced real-time PCR technique represents the method of choice compared to previous, conventional endpoint PCR due to a very sensitive quantification of the viral load over a wide dynamic range (7,12,15,18,20,21,24,31,32,35,41). At present, sample preparation is a major weakness in molecular tests, and improvements are constantly introduced to decrease the variability of the techniques and the risk of contamination, such as ready-to-use reagents and automation of the extraction procedure.A fully automated system, the COBAS AmpliPrep-COBAS TaqMan HBV test (CAP-CTM; Roche Molecular Systems, Inc., Branchburg, NJ) consisting of two integrated platformsthe COBAS AmpliPrep for automated nucleic acid extraction from plasma specimens and the COBAS TaqMan 48, a realtime PCR assay based on TaqMan technology-has recently been developed (13,35). Important improvements compared to other real-time PCR assays for HBV DNA are the incorporation of an internal quantitation standard to monitor the efficiency of the entire process and the introduction of a system to prevent carryover contamination. CAP-CTM is suitable for large routine series and has been demonstrated to equally quantitate HBV genotypes A through G (13, 35), but at present there are only few clinical data (13,28).In the present study, the performance of the CAP-CTM was evaluated and compared to the endpoint PCR assay COBAS AMPLICOR HBV monitor (CAHBM; Roche Molecular Systems). Analytical sensitivity and precision were assessed with an HBV proficiency panel, while correlation and differences in * Corresponding author. Mailing address: Laboratory of Microbiology, Molinette Hospital, Corso Bramante 88/90,

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Section: Discussionsupporting

confidence: 75%

Section: Discussionmentioning

confidence: 99%

COBAS AmpliPrep-COBAS TaqMan Hepatitis B Virus (HBV) Test: a Novel Automated Real-Time PCR Assay for Quantification of HBV DNA in Plasma

et al. 2007

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“…11 In the ongoing trials, we can now use the more recently developed real-time PCR assays (HBV Beacom assay and COBAS TaqMan HBV), which have a dynamic range of 1.0ϫ10 2 to 1.0ϫ10 9 /mL. 12,13 As shown previously in animal models 8 and by a phase I/II short-term clinical trial, 9 clevudine has the unique property of delayed time to viral rebound after treatment. The mechanisms of delayed viral recrudescence observed for 24 weeks after dosing remain to be elucidated.…”

Section: Discussionmentioning

confidence: 99%

A 12-week clevudine therapy showed potent and durable antiviral activity in HBeAg-positive chronic hepatitis B

et al. 2006

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Clevudine is a nucleoside analog with an unnatural ␤-L configuration. In a phase I/II clinical trial, once daily doses ranging from 10 to 200 mg for 28 days were well tolerated, and produced significant antiviral activity. The present study was conducted to assess the degree and durability of the antiviral response to 12 weeks of clevudine treatment, and to investigate its safety and tolerability. A total of 98 patients with HBeAg-positive chronic hepatitis B were randomized to placebo (n ‫؍‬ 32), 30-mg clevudine (n ‫؍‬ 32), and 50-mg clevudine (n ‫؍‬ 34) groups. Patients were followed up after 12 weeks of treatment for a further 24 weeks off-therapy. Median serum hepatitis B virus DNA reductions from baseline at week 12 were 0.20, 4.49, and 4.45 log 10 copies/mL in the placebo, 30-mg clevudine, and 50-mg clevudine groups, respectively (P < .0001). Posttreatment antiviral activities were sustained, with 3.32 and 2.99 log 10 reductions at week 12 off-therapy and 2.28 and 1.40 log 10 reductions at week 24 off-therapies in the 30-and 50-mg clevudine groups, respectively. Median serum alanine aminotransferase (ALT) levels decreased markedly from baseline during clevudine treatment and were maintained below the upper limit of normal throughout the 24 weeks off-therapy in the two clevudine-treated groups. The incidences of adverse events and treatment-emergent grade 3 or 4 laboratory abnormalities were similar for the three groups. In conclusion, clevudine showed potent antiviral activity during therapy and induced a sustained posttreatment antiviral effect for 6 months after a 12-week treatment period, and this was associated with a sustained normalization of ALT levels. (HEPATOLOGY 2006;43:982-988.)

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“…The COBAS TaqMan HBV test (CTM HBV test; Roche Diagnostics, Meylan, France) is a recent commercial nucleic acid amplification test for HBV DNA viral load determination based on TaqMan PCR chemistry (16,23,37).…”

mentioning

confidence: 99%

Evaluation of the COBAS AmpliPrep-Total Nucleic Acid Isolation-COBAS TaqMan Hepatitis B Virus (HBV) Quantitative Test and Comparison to the VERSANT HBV DNA 3.0 Assay

Ronsin¹,

Pillet²,

Bali³

et al. 2006

J Clin Microbiol

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Quantitative detection of hepatitis B virus (HBV) in serum or plasma has become the most direct and reliable method for monitoring chronic hepatitis B. Here, we report the performance characteristics of a real-time PCR hepatitis B DNA quantitative assay, the COBAS TaqMan (CTM) HBV test (Roche Diagnostics, Meylan, France), in combination with an automated DNA extraction on the COBAS AmpliPrep (CAP) instrument using the total nucleic acid isolation kit (TNAI kit), a generic reagent for nucleic acid isolation (both from Roche Diagnostics). The linearity, accuracy, and specificity of the CAP-TNAI-CTM HBV test were evaluated using various reference panels and standards (HBV panel 2004 from Quality Control for Molecular Diagnostics, OptiQuant HBV panel from AcroMetrix, WHO International Standard for HBV, and Teragenix hepatitis B genotype panel). Quantitative results show that the CAP-TNAI-CTM HBV test performed well with respect to linearity, accuracy, and reproducibility from at least 100 to 500,000 HBV DNA IU/ml. Based on the log 10 IU of HBV DNA/ml measured, the intra-assay variation ranged from 2.49% to 8.46% and the interassay variation ranged from 1.88% to 7.83%. The test was extremely sensitive and could detect samples containing HBV DNA below the reported quantification threshold (<30 IU/ml). All HBV genotypes were correctly amplified, and no cross-contamination occurred during the automated sample preparation. In addition, 402 human serum samples were tested comparatively to the VERSANT HBV DNA 3.0 assay (bDNA; Bayer Diagnostics, Puteaux, France). The viral load results of the CAP-TNAI-CTM test and bDNA were significantly correlated, but the agreement between the two tests was poor, with large differences between results for individual samples. The hands-on time was estimated to be reduced from 2.30 h with bDNA to 45 min with the CAP-TNAI-CTM test, and up to 84 samples were completely processed within a working day. Overall, the performance characteristics of the CAP-TNAI-CTM test demonstrated that it provides a high-throughput sensitive and reliable method for quantitation of HBV DNA levels in the routine molecular laboratory.Hepatitis B virus (HBV) is a major causative agent of chronic hepatitis. Approximately 350 million to 400 million people worldwide are chronically infected. Chronic HBV infection can lead to liver cirrhosis and hepatocellular carcinoma (13). Detection and quantification of HBV DNA in serum or plasma are now considered important indicators for managing disease in HBVinfected patients and predicting and monitoring the efficiency of antiviral treatment as well as identifying the emergence of drug resistance by detecting HBV DNA breakthrough (4, 24). Several commercial molecular tests for the quantitation of HBV DNA in serum or plasma have been developed and are routinely used in diagnostic virology laboratories (10, 29). These molecular tests are based on target amplification (HBV Monitor test; Roche Diagnostics, Meylan, France), branched-DNA signal amplification (VERSANT HBV DNA 3.0 assa...

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Dynamic Range and Reproducibility of Hepatitis B Virus (HBV) DNA Detection and Quantification by Cobas Taqman HBV, a Real-Time Semiautomated Assay

Cited by 25 publications

References 25 publications

COBAS AmpliPrep-COBAS TaqMan Hepatitis B Virus (HBV) Test: a Novel Automated Real-Time PCR Assay for Quantification of HBV DNA in Plasma

COBAS AmpliPrep-COBAS TaqMan Hepatitis B Virus (HBV) Test: a Novel Automated Real-Time PCR Assay for Quantification of HBV DNA in Plasma

A 12-week clevudine therapy showed potent and durable antiviral activity in HBeAg-positive chronic hepatitis B

Evaluation of the COBAS AmpliPrep-Total Nucleic Acid Isolation-COBAS TaqMan Hepatitis B Virus (HBV) Quantitative Test and Comparison to the VERSANT HBV DNA 3.0 Assay

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