Digoxigenin 탐식자를 이용한 혈청내 B형간염 바이러스 DNA의 검출

Abstract

To estimate the clinical efficiency of digoxigenin (DIG) labeled DNA probe for the detection of hepatitis B virus (HBV) DNA, 149 HBs Ag positive and 15 normal serum samples were tested by DNA dot blot hybridization using 32P-dCTP-labeled HBV DNA probe and DIG-dUTP-Iabeled HBV DNA probe. Serological markers were tested by ELISA. The sensitivity of 32P-dCTP-labeled HBV DNA probe in detecting HBV DNA was 0.5μg, that of DIG-dUTP-labeled HBV DNA probe labeled by random primed labeling method was 4μg, and the sensitivity of DIG-dUTP-labeled HBV DNA probe labeled by polymerase chain reaction (PCE) was 0.5μg Expire time of 32P-dCTP-Iabeled HBV DNA probe was within a week, that of DIG-dUTP-labeled HBV DNA probe labeled by random primed labeling method was 4 to 6 months, and that of DIG- dUTP-labeled HBV DNA probe labeled by PCR was 2 to 4 weeks. Time needed for autoradiography was 24-48 hours, but in the case of DIG-labeled probe, visualization of the result needed only 6 hours.

The relationship between serum HBV DNA positivity by DIG-labeled probe and serological mark¬ers positivity was evaluated. Twenty-seven(79.4%) of 34 patients positive for both HBs Ag and HBs Ag were positive for HBV DNA compared with 25(21.7%) of 115 HBs Ag positive but HBs Ag negative individuals. There was a statistically significant correlation between the positivity of HBV DNA in serum and the presence or absence of HBe Ag. None of the 15 HBs Ag and HBe Ag negative individuals tested was positive for HBV DNA. But the detection rate of HBV DNA in the cases of HBe Ag negative was as high as 21.7%(25/115), so we concluded that dot blot hybridization technic using DIG-labeled HBV DNA probe could be very efficiently applied for both diagnosing HBV infection and evaluating the infectivity of the HBs Ag positive sera. Key Words：Hepatitis B virus, Dot blot hybridization