Evaluation of the Epstein-Barr Virus R-Gene Quantification Kit in Whole Blood with Different Extraction Methods and PCR Platforms

Fafi‐Kremer, Samira; Morand, Patrice; Barranger, C.; Barguès, Gérard; Magro, Stéphane; Bés, Jérôme; Bourgeois, Philippe; Joannes, Martine; Seigneurin, Jean‐Marie

doi:10.2353/jmoldx.2008.070054

Cited by 24 publications

(23 citation statements)

References 28 publications

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“…The discordant results were observed mostly for EBV loads below 3 log 10 copies/ml, as has been reported previously (1,2,5,12,16). In the transplantation setting, these discrepancies among low EBV loads in whole blood are most often irrelevant for the diagnosis of posttransplantation lymphoproliferative disorders (16).…”

Section: Discussionsupporting

confidence: 49%

“…The laboratory-developed PCR and the Argene PCR assays, described elsewhere (3,5), targeted the BXLF1 thymidine kinase gene and were run on a LightCycler platform (version 2.0) and the Rotor-Gene 6000 platform, respectively. The Artus PCR targeted the EBNA1 gene and was run on the Rotor-Gene 6000 platform.…”

Section: Methodsmentioning

confidence: 99%

“…This heterogeneity is a barrier to the optimal use of quantitative PCR assays. The automation of nucleic acid extraction procedures and the use of commercialized PCR assays could decrease this heterogeneity and improve the agreement and the clinical utility of EBV load measurements in routine settings (5,15,17).…”

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confidence: 99%

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Comparison of Commercial Extraction Systems and PCR Assays for Quantification of Epstein-Barr Virus DNA Load in Whole Blood

et al. 2012

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bThe automation of DNA extraction and the use of commercial quantitative real-time PCR assays could help obtain more reliable results for the quantification of Epstein-Barr virus DNA loads (EBV VL). This study compared two automated extraction platforms and two commercial PCRs for measurement of EBV VL in 10 EBV specimens from Quality Control for Molecular Diagnostics (QCMD) and in 200 whole-blood (WB) specimens from transplant (n ‫؍‬ 137) and nontransplant (n ‫؍‬ 63) patients. The WB specimens were extracted using the QIAcube or MagNA Pure instrument; VL were quantified with the EBV R-gene quantification kit (Argene) or the artus EBV RG PCR kit (Qiagen) on the Rotor-Gene 6000 real-time analyzer; and the results were compared with those of a laboratory-developed PCR. DNA was extracted from the QCMD specimens by use of the QIAamp DNA minikit and was quantified by the three PCR assays. The extraction platforms and the PCR assays showed good correlation (R, >0.9; P, <0.0001), but as many as 10% discordant results were observed, mostly for low viral loads (<3 log 10 copies/ml), and standard deviations reached as high as 0.49 log 10 copy/ml. In WB but not in QCMD samples, Argene PCR tended to give higher VL values than artus PCR or the laboratory-developed PCR (mean difference for the 200 WB VL, ؊0.42 or ؊0.36, respectively). In conclusion, the two automated extraction platforms and the two PCRs provided reliable and comparable VL results, but differences greater than 0.5 log 10 copy/ml remained between the two commercial PCRs after common DNA extraction.

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Section: Discussionsupporting

confidence: 49%

Section: Methodsmentioning

confidence: 99%

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Comparison of Commercial Extraction Systems and PCR Assays for Quantification of Epstein-Barr Virus DNA Load in Whole Blood

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“…Chelating agent Urine these problems is by engaging diagnostic laboratories employing molecular-based diagnostics in well-defined and robust qualitycontrol programs [34]. This is a technique that has worked well in standardizing the detection of influenza in avian samples among the EU and the World Organisation for Animal Health reference laboratories.…”

Section: Potential Problems In Developing Molecular Detection Methodsmentioning

confidence: 99%

“…This creates problems in that diagnostic laboratories may be using different assays with differing degrees of sensitivity and specificity. Indeed, it has been shown that the use of different in-house real-time PCR tests can impair direct comparison of diagnostics between laboratories [34], One key way of subverting…”

Section: Potential Problems In Developing Molecular Detection Methodsmentioning

confidence: 99%

Development of rapid, automated diagnostics for infectious disease: advances and challenges

Ince

McNally²

2009

Expert Review of Medical Devices

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The last 2 years has seen an exponential rise in the amount of research funding made available for the development of rapid diagnostic devices for infectious agents of medical importance. This review reports on several such projects. These highlight the development of fully automated devices for rapid diagnostics, ranging from fully automated real-time PCR-based detection methods to fully automated PCR-and array-based machines for the detection and typing of influenza. This review will also highlight the importance of refocusing work on classical immunoassay techniques, showing how biosensor-based immunoassays can greatly enhance existing assays and at a much reduced cost to molecular-based methods.

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Molecular Detection of Herpesviruses

Kessler

Heyszl

2010

Diagnostic Virology Protocols

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In routine molecular diagnostics, detection of herpesviruses has made a major impact. Infection with herpesviruses is indicated by demonstrating the presence of the virus in selected specimens. Rapid and reliable detection of herpesvirus DNA helps to decrease the lethality as well as the sequelae of herpesvirus infection in patients at risk. This chapter discusses specimen types and both laboratory-developed and commercially available assays useful for molecular detection of herpesviruses. To meet the need for reliable laboratory results, it is advisable to employ maximum automated and standardized kits based on reagents and standards of reproducible high quality. In the routine diagnostic laboratory, introduction of IVD/CE and/or FDA-labeled tests is preferred.

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Evaluation of the Epstein-Barr Virus R-Gene Quantification Kit in Whole Blood with Different Extraction Methods and PCR Platforms

Cited by 24 publications

References 28 publications

Comparison of Commercial Extraction Systems and PCR Assays for Quantification of Epstein-Barr Virus DNA Load in Whole Blood

Comparison of Commercial Extraction Systems and PCR Assays for Quantification of Epstein-Barr Virus DNA Load in Whole Blood

Development of rapid, automated diagnostics for infectious disease: advances and challenges

Molecular Detection of Herpesviruses

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