title: A latest and promising approach for prediction of viral load in hepatitis B virus infected patients.
abstract: BACKGROUND
  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.
  
  
  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.
  
  
  CONCLUSIONS
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Department of Gastroenterology, Centre for Liver Research and Diagnostics, Owaisi Hospital and Research Centre, Kanchanbagh, Hyderabad, India.
authors.forename: Naresh
authors.initials: N
authors.lastname: Yalamanchili
authors.forename: Rahamathullah
authors.initials: R
authors.lastname: Syed
authors.forename: Madhavi
authors.initials: M
authors.lastname: Chandra
authors.forename: Vishnupriya
authors.initials: V
authors.lastname: Satti
authors.forename: Ramachandra
authors.initials: R
authors.lastname: Rao
authors.forename: Aejaz Habeeb
authors.initials: AH
authors.lastname: Mohammed
authors.forename: Khaja Mohammed
authors.initials: KM
authors.lastname: Nanne