Real-Time Polymerase Chain Reaction-Based Detection and Quantification of Hepatitis B Virus DNA

Satyabrata Bag; Santanu Ghosh; Roopa Lalwani; Pratishtha Vishnoi; Sudheer Gupta; Dhirendra Dubey

doi:10.3791/66249

B型肝炎ウイルスDNAのリアルタイムポリメラーゼ連鎖反応に基づく検出と定量(英語)

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Real-Time Polymerase Chain Reaction-Based Detection and Quantification of Hepatitis B Virus DNA

B型肝炎ウイルスDNAのリアルタイムポリメラーゼ連鎖反応に基づく検出と定量(英語)

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04:11 min

December 15, 2023

DOI: 10.3791/66249-v

Satyabrata Bag¹, Santanu Ghosh¹, Roopa Lalwani¹, Pratishtha Vishnoi¹, Sudheer Gupta¹, Dhirendra Dubey¹

¹3B BlackBio Biotech India Limited

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Overview

This study presents a real-time PCR-based protocol for the detection and quantification of hepatitis B virus (HBV) DNA, which is essential for accurate diagnosis and monitoring of HBV infections. The method is recognized as the gold standard due to its sensitivity and specificity in identifying HBV viral loads.

Key Study Components

Research Area

Hepatitis B virus (HBV) detection
Viral load quantification
Clinical diagnostics

Background

Importance of accurate HBV detection in clinical settings
Challenges in identifying low viral loads and occult infections
Comparison of PCR with other diagnostic techniques

Methods Used

Real-time polymerase chain reaction (PCR)
Samples include HBV positive controls
Use of WHO international standard for calibration

Main Results

Successful quantification of HBV DNA using a standard curve
Identification of samples with significant amplification
Parameter calibration ensuring high accuracy and reproducibility

Conclusions

This study provides a reliable protocol for HBV DNA detection and viral load measurement.
The method’s high sensitivity makes it a valuable tool in hepatitis research and clinical diagnostics.

Frequently Asked Questions

What is the significance of detecting HBV DNA?

Detecting HBV DNA is crucial for diagnosing HBV infections and monitoring treatment effectiveness.

How does real-time PCR work?

Real-time PCR amplifies DNA and simultaneously quantifies it using fluorescent dyes during the PCR cycle.

What challenges exist in HBV detection?

Challenges include detecting low viral loads and accommodating various HBV genotypes.

How is the accuracy of PCR results ensured?

Accuracy is ensured by standardizing threshold settings across samples and validating with controls.

What role do quantification standards play?

Quantification standards provide a reference for measuring unknown samples, ensuring reliable results.

Can this protocol detect new mutations?

While the protocol is sensitive, emerging mutations may still pose challenges in accurate detection.

Is this method suitable for clinical use?

Yes, this real-time PCR method is fast, sensitive, and suited for clinical diagnostics.

B型肝炎ウイルス(HBV)DNAのリアルタイムポリメラーゼ連鎖反応(PCR)ベースの検出および定量は、HBV感染を診断およびモニタリングするための高感度で正確な方法です。ここでは、HBV DNA検出とサンプルのウイルス量測定のプロトコルを紹介します。

このビデオでは、PCR HBVウイルス量キットを介して、体外診断用IVD承認のリアルタイムPCR廃棄物を使用して、HBV DNAを検出し、そのウイルス量を定量するためのステップバイステップのプロトコルを紹介します。HBV診断研究は、リアルタイムPCR、デジタルPCR、NGS、血清学的検査を使用します。リアルタイムPCRは、HBV DNA検出とウイルス量定量のゴールドスタンダードです。

デジタルPCRは、ウイルスの絶対量を定量化します。NGSは新しい変異を特定し、血清学的検査は抗原または抗体を検出します。低ウイルス量症例と潜伏性HBV感染の検出には、いくつかの実験上の課題があります。

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