Simultaneous detection of Hepatitis B virus and Hepatitis C virus in human plasma using Taq-man chemistry
Journal Title: Journal of Medical and Allied Sciences - Year 2011, Vol 1, Issue 2
Abstract
Designing a rapid, reliable and sensitive assay, for detection of hepatitis B virus and Hepatitis C virus variants by real-time PCR, is challenging at best. A recent approach for quantifying the viral load using the sensitive fluorescence principle, was used in this study. A total of 350 samples were collected from outpatient unit, CLRD. Complete Human HBV DNA and HCV sequences were obtained from the National Centre for Biotechnology Information (NCBI); primers and probes were designed and synthesized from core, surface and x region of Hepatitis B and UTR region of HCV. Real-time based detection was done, using standard kit and in-house generated standards and RT-PCR protocols. A standard curve was generated by using the Smart Cycler II software and serial dilution 102 to 108 of cloned viral regions, the calibration curve was linear in a range from 102 to108 cp/ml for both HBV and HCV, with R2 value of 0.999 and 0.995. Out of 100 predetermined HCV negative samples, 02 samples were found positive with in-house developed RTPCR assay, the positivity of this sample was confirmed by sequencing the amplified product. Low cost of this assay procedure and précised sample volume will permit the assay to be implemented for routine screening of Hepatitis B and C virus mono-infection and co-infection using Real Time PCR , Nucleic acid Chip technology and Fluorescent End Point detection systems. 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 HBV and HCV virus monitor.
Authors and Affiliations
Naresh Kumar Y| Center for Liver Research and Diagnostics, Deccan College of Medical Sciences and Allied Hospitals, Kanchanbagh, Hyderabad-500058, Andhra Pradesh, India, Rahmathulla S| Center for Liver Research and Diagnostics, Deccan College of Medical Sciences and Allied Hospitals, Kanchanbagh, Hyderabad-500058, Andhra Pradesh, India, Madhavi C| Center for Liver Research and Diagnostics, Deccan College of Medical Sciences and Allied Hospitals, Kanchanbagh, Hyderabad-500058, Andhra Pradesh, India, Ramachandra V V| Center for Liver Research and Diagnostics, Deccan College of Medical Sciences and Allied Hospitals, Kanchanbagh, Hyderabad-500058, Andhra Pradesh, India, Vishnupriya S| Department of Genetics, Osmania University, Tarnaka, Hyderabad-500007, Andhra Pradesh, India, Mohammed Aejaz Habeeb| Center for Liver Research and Diagnostics, Deccan College of Medical Sciences and Allied Hospitals, Kanchanbagh, Hyderabad-500058, Andhra Pradesh, India, Khaja M N. | Center for Liver Research and Diagnostics, Deccan College of Medical Sciences and Allied Hospitals, Kanchanbagh, Hyderabad-500058, Andhra Pradesh, India
Keywords
Related Articles
- EP ID EP12652
- DOI -
- Views 515
- Downloads 52
Comparative study of risk factors and lipid profile pattern in ischemic and haemorrhagic stroke
Stroke is a devastating and disabling cerebrovascular disease with significant amount of residual deficit leading on economic loss. Creating awareness on common risk factors will reduce the incidence of stroke. The stu...
High Mallampati score, obesity and obstructive sleep apnea: Triple insult to lung function?
The paper assesses the combined effect of high Mallampati score, obesity and obstructive sleep apnea (OSA) on lung function as measured by spirometry. Our results showed that the combination of sleep apnea, obesity an...
Estimation of height of an individual from forearm length on the population of Eastern India
Height is a fundamental unit to assess growth and nutrition, for calculating body surface area and predicting pulmonary function in a person. But measurement of height may be hindered by muscle weakness, joint & spina...
Old staining methods: Dyeing a slow death
The book of infectious diseases—a living document