A latest and promising approach for prediction of viral load in hepatitis B virus infected patients
| Autor: | Vishnupriya Satti, Naresh Yalamanchili, Khaja Mohammed Nanne, Aejaz Habeeb Mohammed, Madhavi Chandra, Ramachandra Rao, Rahamathullah Syed |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2011 |
| Předmět: |
Hepatitis B virus
Reproducibility Serial dilution Calibration curve Biology medicine.disease_cause Serum samples Virology Molecular biology quantification real-time Polymerase Chain Reaction Real-time polymerase chain reaction Genetics medicine genetic markers Original Article TaqMan chemistry 19p 13.3 Viral load Genetics (clinical) |
| Zdroj: | Indian Journal of Human Genetics |
| ISSN: | 1998-362X 0971-6866 |
| Popis: | Introduction: Designing a rapid, reliable and sensitive assay for detection of hepatitis B virus (HBV) variants by real-time PCR is challenging at best. A recent approach for quantifying the viral load using a sensitive fluorescent principle was brushed in this study. Materials and Methods : A total of 250 samples were collected from the outpatient unit, CLRD. Complete Human HBVDNA sequences ( n = 944) were selected from the National Centre for Biotechnology Information (NCBI), primers and probes were designed and synthesized from the core, surface, and x region. Real-time based quantification was carried out using a standard kit and in-house generated standards and RT-PCR protocols. Results and Discussion: The standard calibration curve was generated by using serial dilution 10 2 to 10 8 . The calibration curve was linear in a range from 10 2 to 10 8 copies/ml, with an R 2 value of 0.999. Reproducibility as measured by dual testing of triplicates of serum samples was acceptable, with coefficients of variation at 6.5%, 7.5%, and 10.5%. Our results showed that amplification performance was good in the case of the x-region-based design (98%). Out of 100 negative samples screened by enzyme linked immunosorbent assay and the standard RT-PCR kit, one sample was detected as positive with the in-house developed RT-PCR assay, the positivity of the sample was confirmed by sequencing the amplified product, NCBI accession EU684022. Conclusion: This assay is reproducible showing limited inter- and intra-assay variability. We demonstrate that the results of our assay correlated well with the standard kit for the HBV viral load monitor. |
| Databáze: | OpenAIRE |
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