Impact of Hepatitis C Virus (HCV) Genotypes on Quantification of HCV RNA in Serum by COBAS AmpliPrep/COBAS TaqMan HCV Test, Abbott Antiretroviral Therapy HCV RealTime Assay, and VERSANT HCV RNA Assay

Abstract

The VERSANT HCV RNA 3.0 (bDNA), COBAS AmpliPrep/COBAS TaqMan HCV, and Abbott ART HCV RealTime assays were compared for hepatitis C virus RNA quantification in 158 clinical specimens (genotypes 1 to 5). RNA values differed significantly between methods (P < 0.0001), and mean titer differences ranged from 0.01 to 0.50 log 10 IU/ml depending on the genotypes.Hepatitis C virus (HCV) infection is a major cause of chronic liver disease. Therapy is monitored by assessing the kinetics of viral clearance based on HCV RNA quantitation (2,8,12,17). Available commercial kits are based on target amplification technology (reverse transcription-PCR) or on signal amplification (branched DNA). Irrespective of the method, the specificity of the hybridization between probe/primer and target is the determining factor of the method's performance. Polymorphisms within the primer/probe-targeted HCV regions may cause significant losses of performance and compromise the accuracy of viral load (VL) measurements. Based on their natural polymorphism, the HCV viruses have been classified into six major genotypes (GT) (14). Most known GT are found in Europe but with different distribution frequencies in various geographic regions (5).The aim of the present study was to evaluate the performance of the three commercially available HCV RNA quantitative assays, two real-time reverse transcriptase assays, and a signal amplification assay using a panel of serum samples from well-characterized HCV-infected patients with GT1 to GT5.Serum samples were collected during 2005 from 158 consecutive patients residing in Southern France, with HCV infection confirmed by use of a COBAS Amplicor HCV test, version 2.0 (detection limit of 50 IU/ml; Roche Diagnostics, Mannheim, Germany). Each serum sample was divided into three aliquots and stored at Ϫ80°C within 4 h of blood sampling. HCV genotype was determined by using a VERSANT HCV genotyping assay (line probe assay) (Bayer Diagnostics, Tarrytown, NY).All samples were processed in accordance with the manufacturer's instructions. The COBAS Ampliprep/COBAS TaqMan HCV test (CAP/CTM) (Roche Diagnostics, Mannheim, Germany) requires 1,050 l of serum with a sample input volume of 850 l and with a quantification range of 15 to 6.9 ϫ 10 7 IU/ml (1.18 to 7.84 log 10 ). The HCV RealTime assay (antiretroviral therapy [ART]; Abbott Laboratories, Abbott Park, IL) requires 1,000 l of serum, with a quantification range of 12 to 1 ϫ 10 7 IU/ml (1.08 to 7.0 log 10 ). The Abbott ART HCV kit was used in conjunction with the Abbott m1000 extraction and the m2000rt amplification systems. The VERSANT HCV RNA 3.0 (bDNA) assay (Bayer Diagnostics, Tarrytown, NY) requires 50 l of plasma or serum, with a quantification range of 615 to 7.7 ϫ 10 6 IU/ml (2.79 to 6.89 log 10 ) with the VERSANT 340 analyzer.Values outside the quantification ranges were exclude from comparison and correlation analyses. Mean values Ϯ standard deviations (SD) were determined for each method and compared using the Student paired t test for global results. Diffe...

Real-Time PCR Assays for the Quantification of HCV RNA: Concordance, Discrepancies and Implications for Response Guided Therapy

“…Therefore the question arose how to deal with the significantly higher proportion of on-treatment samples with low levels of detectable HCV RNA (<LLOQ) at week 4/8 by the ART assay. As the specificity of the ART assay had already been evaluated in independent performance studies [ 24 – 31 ] and since discrepancies were noted mainly in week 4/8 samples but not in subsequent samples, the probability of ´false positive´ results seems to be very low. In addition a recent study by Wiesmann et al [ 32 ] compared 6 different assays (Roche CAP/CTM HCV Ver.…”

Performance of the Novel QiagenartusQS-RGQ Viral Load Assays Compared to That of the Abbott RealTime System with Genetically Diversified HIV and Hepatitis C Virus Plasma Specimens

“…In contrast, the Abbott RealTime assays were introduced into the market in 2005 and have become known for their good performance on diverse HIV subtypes and HCV genotypes (16,20). In this study, we have compared the quantitative correlations of the Abbott RealTime (Darmstadt, Germany) and Qiagen artus QS-RGQ (Hilden, Germany) viral load assays on a genetically diversified panel of clinical HIV and HCV specimens.…”

“…For HCV, the overall Qiagen performance was good, but the assay systematically yielded higher concentrations in comparison to Abbott RealTime, mainly for genotypes 4 and 5. However, comparable or higher systematic deviations for non-1 HCV genotypes and non-B HIV subtypes between other well-established viral load assays are well known (7,11,15,20,21). Nevertheless, the relative overquantification of HCV needs to be communicated to health care professionals in order to prevent inadequate therapeutic de- cisions.…”

A Correlative Study on Hepatitis C Virus Load Determined by Real Time Polymerase Chain Reaction with Serum Biomarkers in Patients with Renal Disease

“…Several researchers have developed Reverse Transcriptase based Real Time PCR [15][16][17][18][19][20][21][22][23][24], assays for detection and quantitation of HCV RNA. A relationship has been suggested between HCV type, subtype, and serum HCV RNA levels [6].…”