Diagnostic microbiology and infectious disease · 2024
A molecular detection method for HBV pgRNA without RNA extraction based on nucleic acid hybridization.
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Hao Xiao, Yuan Min, Li Yanmei, Li Ruirui, Li Xuejing, Yao Meixue
Аннотация
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.
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.