A latest and promising approach for prediction of viral load in hepatitis B virus infected patients.
Indian J Hum Genet
;
2011 Jan; 17(1): 17-21
Article
in English
| IMSEAR
| ID: sea-138926
ABSTRACT
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 ANDMETHODS:
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 ANDDISCUSSION:
The standard calibration curve was generated by using serial dilution 102 to 108. The calibration curve was linear in a range from 102 to 108 copies/ml, with an R2 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 {"type""entrez-nucleotide","attrs"{"text""EU684022","term_id""189176131"}}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.
Full text:
Available
Index:
IMSEAR (South-East Asia)
Main subject:
Patients
/
Humans
/
Genetic Markers
/
Hepatitis B virus
/
Taq Polymerase
/
Real-Time Polymerase Chain Reaction
/
Hepatitis B
/
India
Type of study:
Prognostic study
Country/Region as subject:
Asia
Language:
English
Journal:
Indian J Hum Genet
Year:
2011
Type:
Article
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