Reverse transcriptase droplet digital PCR vs reverse transcriptase quantitative real-time PCR for serum HBV RNA...

Area of Science:

• Virology
• Molecular Biology
• Hepatology

Background:

• Serum hepatitis B virus (HBV) RNA is a key marker for covalently closed circular DNA activity.
• Detecting HBV RNA presents significant methodological challenges.
• Chronic hepatitis B (CHB) diagnosis and monitoring require accurate viral load assessment.

Purpose of the Study:

• To compare the performance of RT-ddPCR and RT-qPCR for quantifying serum HBV RNA.
• To evaluate HBV RNA levels in treatment-naïve CHB patients across different disease phases.
• To determine the optimal method for HBV RNA quantification in clinical settings.

Main Methods:

• Serum samples from 417 treatment-naïve CHB patients (136 HBeAg-positive, 281 HBeAg-negative) were analyzed.
• HBV RNA was quantified using both RT-ddPCR and RT-qPCR assays.
• Assay performance was evaluated based on linearity, quantitative correlation, limit of detection, and consistency.

Main Results:

• RT-ddPCR and RT-qPCR showed high linearity and quantitative correlation for HBV RNA measurement.
• RT-ddPCR demonstrated a lower limit of detection (10^2 copies/mL) than RT-qPCR (10^3 copies/mL).
• RT-ddPCR exhibited superior consistency in quantifying HBV RNA across all concentrations and enhanced sensitivity in HBeAg-negative patients.

Conclusions:

• RT-ddPCR is a more sensitive and consistent method for serum HBV RNA quantification than RT-qPCR.
• This technique is particularly beneficial for detecting HBV RNA in HBeAg-negative CHB patients with low viral loads.
• RT-ddPCR represents an optimal method for clinical HBV RNA quantification.