DIRECT-DETECTION OF HEPATITIS-B VIRUS GENE INTEGRATED IN THE ALEXANDER CELL USING FLUORESCENCE IN-SITU POLYMERASE CHAIN-REACTION

Prolonged infection of hepatitis B virus (HBV) is reported to cause he patocellular carcinoma (HCC) via liver cirrhosis, However, it is still unknown whether the HCC is induced by the HBV DNA integration or by i nflammatory stimulation during the phase of liver cirrhosis, The aim o f this study is to determine the intracellular or intranuclear distrib ution of HBV DNA with a highly sensitive assay. Here we directly detec ted the integration of HBV DNA by fluorescence in situ polymerase chai n reaction (FISPCR). Since FISPCR products directly incorporate rhodam iile-4-dUTP, the nucleus of Alexander cells integrated with HBV gene r eacted with the HBV primers emits obvious fluorescence. The fluorescen ce values which were measured with an imaging analyzer show a signific ant difference between Alexander cells as compared to the controls. In conclusion, the target sequences of HBV were specifically amplified a s fluorescent DNA after the present FISPCR procedure. This method coul d provide a novel and simple strategy for determining the quantitative role of viral DNA integration in oncogenesis. (C) 1997 Elsevier Scien ce Ireland Ltd.