[Loop-mediated isothermal amplification for visual detection of hepatitis B virus]

Area of Science:

• Molecular Biology
• Virology
• Biotechnology

Background:

• Hepatitis B virus (HBV) infection remains a significant global health concern.
• Accurate and accessible diagnostic methods are crucial for effective HBV management.
• Loop-mediated isothermal amplification (LAMP) is a promising nucleic acid amplification technique.

Purpose of the Study:

• To evaluate the efficacy of loop-mediated isothermal amplification (LAMP) for visual detection of HBV DNA.
• To assess the use of boiled serum as a sample preparation method for LAMP.
• To compare the performance of LAMP with conventional PCR and fluorescent quantitative PCR (FQ-PCR).

Main Methods:

• Designed specific LAMP primers targeting conserved regions of the HBV S gene.
• Optimized reaction conditions for LAMP, comparing DNA extraction using kit and boiling methods.
• Evaluated specificity, sensitivity, and anti-interference capabilities of LAMP using standard strains and clinical samples.
• Visual detection of LAMP results using Hydroxynaphthol blue (HNB) and comparison with electrophoresis and SYBR Green I.

Main Results:

• Optimized LAMP conditions yielded high specificity with no non-specific amplification.
• LAMP demonstrated a sensitivity of 10 copies/tube, irrespective of DNA extraction method.
• HNB visual detection showed comparable sensitivity to electrophoresis and SYBR Green I, with reduced aerosol risk.
• LAMP using boiled serum showed good consistency with FQ-PCR (Kappa = 0.762), outperforming standard PCR (Kappa = 0.186).

Conclusions:

• LAMP offers significant advantages over conventional PCR for HBV detection.
• The visual detection capability of LAMP, combined with boiled serum preparation, simplifies HBV diagnosis.
• LAMP is a viable tool for HBV detection in resource-limited settings, such as primary hospitals and field applications.