We use cookies to understand how you use our site and to improve your experience. This includes personalizing content and advertising. To learn more, click here. By continuing to use our site, you accept our use of cookies. Cookie Policy.

Features Partner Sites Information LinkXpress
Sign In
Advertise with Us
RANDOX LABORATORIES

Download Mobile App




New Tool Developed for Diagnosis of Chronic HBV Infection

By LabMedica International staff writers
Posted on 23 Oct 2019
Print article
Image: Diagram workflow for co-amplification at lower denaturation temperature PCR (COLD-PCR)/fluorescence melting curve analysis (FMCA) (Photo courtesy of The First Affiliated Hospital of Fujian Medical University).
Image: Diagram workflow for co-amplification at lower denaturation temperature PCR (COLD-PCR)/fluorescence melting curve analysis (FMCA) (Photo courtesy of The First Affiliated Hospital of Fujian Medical University).
A team of Chinese researchers has described the development of a new and powerful tool for the diagnosis and treatment of chronic hepatitis B virus (HBV) infection.

Methods currently used to diagnose and monitor chronic hepatitis B (CHB) infection are for the most part based on dynamic and real-time HBV DNA, genotype, and reverse transcriptase (RT) mutation analysis. However, the methods used to perform these analyses are limited by poor sensitivity or inability to detect more than one mutation at a time. Other methods are too cumbersome or expensive for clinical use.

Investigators at Fujian Medical University (Peoples Republic of China) have improved this situation by establishing a highly sensitive co-amplification at lower denaturation temperature PCR (COLD-PCR) coupled with probe-based fluorescence melting curve analysis (FMCA) for precision diagnosis of CHB patients.

COLD-PCR (co-amplification at lower denaturation temperature-PCR) is a modified Polymerase Chain Reaction (PCR) protocol that enriches variant alleles from a mixture of wildtype and mutation-containing DNA. The ability to preferentially amplify and identify minority alleles and low-level somatic DNA mutations in the presence of excess wildtype alleles is useful for the detection of mutations.

Melting curve analysis is an assessment of the dissociation characteristics of double-stranded DNA during heating. As the temperature is raised, the double strand begins to dissociate leading to a rise in the absorbance intensity, hyperchromicity. The temperature at which 50% of DNA is denatured is known as the melting temperature. The information gathered can be used to infer the presence and identity of single-nucleotide polymorphism (SNP) mutations.

The COLD-PCR/FMCA method was shown to be able to detect HBV DNA, genotypes, and RT mutations, simultaneously. The analytical performance of this method, including imprecision, accuracy, sensitivity, detection limits, linear range, and its ability to detect minor variants was systematically evaluated.

Results revealed that the COLD-PCR/FMCA method could detect HBV mutations at much lower concentrations than other techniques such as PCR/FMCA or PCR Sanger sequencing (1% vs. 10% vs. 20%, respectively). The new technique could also distinguish different phases of HBV infection according to the proportion and type of mutations as well as by detecting HBV DNA.

"Guidelines have confirmed that dynamic monitoring of HBV DNA, genotypes, and reverse transcriptase (RT) mutant DNA is of great importance to assess infection status, predict disease progression, and judge treatment efficacy in HBV-infected patients," senior author Dr. Qishui Ou, a researcher in laboratory medicine at The First Afilliated Hospital of Fujian Medical University. "We believe COLD-PCR/FMCA provides a powerful laboratory tool for precise diagnosis and treatment of HBV-infected patients."

"Our goal was to establish a more practical and inexpensive method with high sensitivity to detect genotype and RT mutations while detecting HBV DNA," said Dr. Ou. "Until now there have not been high-throughput approaches to detect HBV DNA, genotype, and RT mutations simultaneously. Therefore, it is necessary to establish a more practical and inexpensive method with high sensitivity to detect genotype and RT mutations while detecting HBV DNA. COLD-PCR/FMCA has that potential."

The study was published in the October 10, 2019, online edition of the Journal of Molecular Diagnostics.

Related Links:
Fujian Medical University


Print article

Channels

Molecular Diagnostics

view channel
Image: Blood smear from a P. falciparum culture: several red blood cells have ring stages inside them while close to the center is a schizont and on the left a trophozoite (Photo courtesy of Wikimedia Commons)

Panel of MicroRNAs Differentiates Uncomplicated and Severe Malaria in Children

MicroRNAs (miRNAs), which are rapidly released from damaged tissues into the host fluids, constitute a promising biomarker for the prognosis of severe malaria. MiRNAs comprise a class of about 20 n... Read more

Pathology

view channel
Image: The Leica Bond III stainer is fully automated Immunohistochemical and In Situ Hybridization (IHC and ISH stainer) (Photo courtesy of Leica Biosystems).

Mismatch Repair/Microsatellite Instability Evaluated Using Cytology Effusion Specimens

DNA mismatch repair (MMR) status is routinely assessed in colorectal and endometrial carcinoma as a method of cancer prevention, surveillance in patients with Lynch syndrome and their families, and for... Read more

Industry

view channel
Image: BioProfile FLEX2 Cell Culture Analyzer (Photo courtesy of Nova Biomedical)

Nova Biomedical Adds Sample Retain Collector to BioProfile FLEX2 Cell Culture Analyzer

Nova Biomedical (Waltham, MA, USA) has added a Sample Retain Collector (SRC) for its BioProfile FLEX2 automated cell culture analyzer which measures up to 16 tests including pH, gases, metabolites, osmolality,... Read more
Copyright © 2000-2021 Globetech Media. All rights reserved.