Assessment of a method for detecting serum HBV DNA with HBV DNA probe labelled directly by alkaline phosphatase.
OBJECTIVE
To assess a sensitive and specific technique for detecting serum HBV DNA with an HBV DNA probe labelled directly by alkaline phosphatase (AlkPhos Direc probe).


METHODS
AlkPhos Direc probe was prepared with purified HBV DNA labelled directly by alkaline phosphatase. The probe and chemiluminescent substrate CDP-star for AP were used in hybridization assay. HBV DNA was detected by autoradiography on a film. The results of 80 samples were compared between the chemiluminescent dot blot hybridization assay with the AlkPhos Direc probe and another assay with the digoxigenin-labelled HBV DNA probe. The correlation of seventy-sample results of fluorescent quantitative HBV DNA PCR assay and dot blot hybridization assay with the AlkPhos Direc probe was analysed.


RESULTS
The sensitivity of the AlkPhos Direc probe was 10 pg at least. The coincidence of the AlkPhos Direc probe was 100% compared with that of the digoxigenin-labelled HBV DNA probe. A correlation coefficient of HBV DNA quantitative results between fluorescent quantitative HBV DNA PCR assay and dot blot hybridization assay with the AlkPhos Direc probe was 0.98 (P<0.01).


CONCLUSIONS
The method detecting HBV DNA in serum with the HBV DNA AlkPhos Direc probe is sensitive and specific. The results of the two assays with the AlkPhos Direc probe or with the digoxigenin-labelled HBV DNA probe are completely coincident. The correlation of HBV DNA quantitative results between fluorescent QPCR assay and dot blot hybridization assay with the AlkPhos Direc probe is satisfactory.
Chen
Ya-Xi
YX
Institute for Viral Hepatitis, Chongqing University of Medical Sciences, Chong-qing 400010, China. zlcyxi@sina.com
Huang
Ai-Long
AL
Qi
Zhen-Yuan
ZY
Shan
You-Lan
YL
Sun
Hang
H
D000469:Alkaline Phosphatase; D015139:Blotting, Southern; D015342:DNA Probes; D004279:DNA, Viral / Q000032:analysis*; D006509:Hepatitis B / Q000175:diagnosis; D006515:Hepatitis B virus / Q000302:isolation & purification*; D006801:Humans; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D015342:DNA Probes; D004279:DNA, Viral; D000469:Alkaline Phosphatase