Comparison of four methods for quantitative measurement of hepatitis B viral DNA

Butterworth, L. A.; Prior, Sharon; J, Buda Phillip; Faoagali, Joan; Cooksley, W. G. E.; Cooksley, E.

doi:10.1016/s0168-8278(96)80264-9

Cited by 48 publications

(40 citation statements)

References 6 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…In the present study, the quantitative analysis showed variations ranging from 6 to 35% (percent positive results outside the defined range) in the actual copy numbers assigned to samples. Similar method-related deviations in quantitation have been reported by others (3,5) In 1999, the Eurohep Pathobiology Group established two international reference plasma preparations, each containing approximately 2.6 ϫ 10 9 copies/ml, thereafter defined as 10 9 Eurohep units (6). These Eurohep samples have been used for the evaluation of commercial kits (10,14), and one of these may be the basis of a World Health Organization reference sample.…”

Section: Discussionsupporting

confidence: 58%

European Proficiency Testing Program for Molecular Detection and Quantitation of Hepatitis B Virus DNA

Valentine-Thon¹,

Loon

Schirm³

et al. 2001

J Clin Microbiol

View full text Add to dashboard Cite

These results demonstrate that while the qualitative performance of HBV detection has considerably improved compared to that of a previously published HBV proficiency study, the detection levels of many commercial quantitative assays are still too high to allow adequate quantitation of all relevant clinical samples.Direct detection and quantitation of hepatitis B virus (HBV) DNA in plasma or serum are now used routinely to evaluate viremia in HBV-infected persons, to identify infectious chronic carriers, and to predict and monitor the efficacy of antiviral therapy (2,8,11). Since the early 1980s, a variety of molecular detection and quantitation methods have been developed, including dot and slot blot hybridization with radioactive and nonradioactive DNA probes (19-21), chemiluminescent detection of HBV DNA-RNA hybrids (14), PCR amplification of HBV DNA followed by hybridization to probes bound to a microwell plate (10, 12, 22) or magnetic beads (13), branched DNA (bDNA) signal amplification of an HBV DNA-DNA hybrid (7), transcription-mediated amplification (9), and fluorescent real-time detection of amplified HBV DNA (1). Each method, calibrated uniquely, exhibits its own sensitivity, specificity, and dynamic range. Standardization is ongoing (5, 6).To assess the relative value of these methods in detecting and quantitating HBV DNA, international proficiency studies with well-characterized, simulated clinical samples would be required. In the first and only such study published to date (17), 39 laboratories analyzed 22 samples, including 12 undiluted samples with and without HBV DNA. (The lowest positive sample contained 3.5 pg/ml, or approximately 980,000 copies/ml.) Only 27.9% of the data sets had all 12 samples correct, and 34.9% showed false-positive results. Clearly, a majority of the participating laboratories had problems with both sensitivity and specificity.The present report describes two recent HBV proficiency panels (lowest viral load of 1,000 copies/ml) designed by the European Union Concerted Action on Quality Control (EU QCCA) of Nucleic Acid Amplification in Diagnostic Virology and prepared by Boston Biomedica, Inc. (BBI; West Bridgewater, Mass.). The results obtained with these panels demonstrate that while the qualitative detection of HBV DNA has significantly improved, the detection levels of many commercial quantitative assays are still too high to allow adequate quantitation of the clinical samples seen in routine diagnostic laboratories. MATERIALS AND METHODSPanels. (i) Preparation. Panels were prepared by BBI from human plasma containing HBV DNA of subtype ad or ay by appropriate dilution in sterile filtered defibrinated plasma (Basematrix) with 0.09% sodium azide as preservative in accordance with the ISO 9001 Quality System Standards and the 21CFR 820 "Good Manufacturing Practice for Medical Devices: General." Plasma units were obtained from Food and Drug Administration-licensed facilities that comply with the applicable federal regulations (21CFR, part 600).The pilot dilutions made for...

show abstract

Section: Discussionsupporting

confidence: 58%

European Proficiency Testing Program for Molecular Detection and Quantitation of Hepatitis B Virus DNA

Valentine-Thon¹,

Loon

Schirm³

et al. 2001

J Clin Microbiol

View full text Add to dashboard Cite

show abstract

“…However, their quantification methodology was based on the solution hybridization assay (Abbott), with a detection limit of 1.6 to 2.0 pg/ml, corresponding to 4.5x10 5 to 5.6x10 5 genomes/ml 4,6,20,31 . Concentrations calculated by the Abbott assay were approximately 40 to 100 fold and 35 fold lower than those obtained when the same sample was tested by the Quantiplex and Hybrid Capture System HBV-DNA assays, respectively 4,17,31 . On the other hand, like us, LAU et al observed a lower rate of response when using a quantitative PCR method 21 .…”

Section: Discussionmentioning

confidence: 99%

Predictive factors for response to Lamivudine in chronic hepatitis B

Silva

Fonseca

Carrilho

et al. 2000

Rev. Inst. Med. trop. S. Paulo

View full text Add to dashboard Cite

show abstract

“…However, the differential diagnosis between patients with HBeAg‐negative CHB and inactive HBsAg carriers on the basis of detectable and undetectable serum HBV DNA is not always simple. The detection of HBV viraemia certainly depends on the sensitivity of the method used [41,42]. Serum HBV DNA is undetectable by molecular hybridization assays in practically all inactive HBsAg carriers and in a proportion of patients with HBeAg‐negative CHB [10,43].…”

Section: Diagnosis Of Pre‐core Hbv Mutant Related Chronic Liver Diseasementioning

confidence: 99%

Diagnosis and management of pre‐core mutant chronic hepatitis B

Papatheodoridis

Hadziyannis

2001

Journal of Viral Hepatitis

102

View full text Add to dashboard Cite

Chronic hepatitis due to pre-core hepatitis B virus (HBV) mutants presents as hepatitis B e antigen (HBeAg)-negative chronic hepatitis B (CHB). HBeAg-negative CHB represents a late phase in the natural course of chronic HBV infection that develops after HBeAg loss and seroconversion to anti-HBe. It is usually associated with pre-core stop codon mutation at nucleotide 1896 (mainly selected in non-A HBV genotypes), but also with other pre-core changes or with mutations in the basic core promoter region (mainly in HBV genotype A). In chronic HBV infections, pre-core mutants can be detected both in patients with HBeAg-negative CHB and in inactive hepatitis B surface antigen (HBsAg) carriers. The diagnosis of HBeAg-negative CHB is based on HBsAg positivity, HBeAg negativity, and mainly on increased alanine aminotransferase (ALT) and serum HBV-DNA levels and exclusion of other causes of liver disease. The differential diagnosis between patients with CHB and inactive HBsAg carriers can be made only by close follow-up of aminotransferase activity and viraemia levels, although the cut-off level of serum HBV DNA has not been definitely determined. IgM anti-HBc levels have also been suggested as an index that increases the diagnostic accuracy for transient hepatitis flares, while liver biopsy confirms the diagnosis and evaluates the severity of the liver disease. Interferon-alpha (IFN-alpha) and lamivudine are the two drugs that have been tried, mainly in the management of HBeAg-negative CHB. A 12-month course of IFN-alpha achieves sustained biochemical remission in about 20% of patients, which has been associated with improvement in the long-term outcome of this subset. A 12-month course of lamivudine is rather ineffective, maintaining remission in less than 15% of patients after cessation of therapy. Long-term lamivudine is associated with progressively increasing rate of virological and subsequent biochemical breakthroughs due to YMDD mutants, with approximately 30% of patients remaining in remission in the third year of therapy. Several other antiviral agents are currently being evaluated in this setting with combined regimens being the most reasonable step for the near future.

show abstract

Comparison of four methods for quantitative measurement of hepatitis B viral DNA

Cited by 48 publications

References 6 publications

European Proficiency Testing Program for Molecular Detection and Quantitation of Hepatitis B Virus DNA

European Proficiency Testing Program for Molecular Detection and Quantitation of Hepatitis B Virus DNA

Predictive factors for response to Lamivudine in chronic hepatitis B

Diagnosis and management of pre‐core mutant chronic hepatitis B

Contact Info

Product

Resources

About