A molecular detection method for HBV pgRNA without RNA extraction based on nucleic acid hybridization

Abstract

Background

Early diagnosis is critical for patients with chronic hepatitis B. Here, we utilized a sandwich RNA hybridization assay to directly detect HBV pgRNA, thereby avoiding the need for RNA extraction and purification.

Methods

We designed a hybridization cascade reaction on a solid surface using a set of oligonucleotide probes that target several highly conserved regions in pgRNA. The detection performance was validated by concurrently testing serum samples from CHB patients and healthy individuals.

Results

The optimal detection conditions were: a universal probe coating concentration of 0.003 µg/µl with a coating duration of 2 h; capture probe and detection probe concentrations of 0.1 nM; a hybridization capture duration of 4 h; and an antibody incubation duration of 60 minutes. The sensitivity and specificity were calculated to be 91.47 % and 90.63 %, respectively.

Conclusion

This novel detection method is both simple and high-throughput, making it particularly suitable for active CHB infection screening