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Detection of HBV‐DNA by in situ hybridization using a biotin‐labeled probe
Author(s) -
Negro Francesco,
Berninger Mark,
Chiaberge Elisabetta,
Gugliotta Patrizia,
Bussolati Giovanni,
Actis Giovanni C.,
Riuetto Mario,
Bonino Ferruccio
Publication year - 1985
Publication title -
journal of medical virology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.782
H-Index - 121
eISSN - 1096-9071
pISSN - 0146-6615
DOI - 10.1002/jmv.1890150407
Subject(s) - in situ hybridization , microbiology and biotechnology , hbcag , hybridization probe , hepatitis b virus , biology , biotinylation , hbsag , virology , southern blot , nucleic acid thermodynamics , dna , immunofluorescence , hepadnaviridae , nucleic acid , biotin , virus , antibody , biochemistry , messenger rna , gene , immunology , base sequence
Abstract A biotin‐labeled DNA probe specific for hepatitis B virus (HBV) nucleotide sequences was hybridized in situ to liver tissue of 20 patients; 16 were chronic carriers of hepatitis B surface antigen (HBsAg) and 4 had no markers of HBV infection. HBV‐DNA was also analyzed in the serum and the liver of these patients by spot and Southern blot hybridization, respectively. Liver specimens from six carriers were positive for HBV‐DNA both by in situ and Southem blot hybridization; ten carriers were negative by in situ hybridiza‐ tion, and two of these were positive by Southern blot technique. The staining was granular, mainly cytoplasmic, limited to liver specimens containing replicative forms of HBV‐DNA, and associated with detection of HBcAg in hepatocytes by immunofluorescence. The sensitivity of this technique was not sufficient to detect few copies of integrated HBV‐DNA. The hybridization procedure was specific, as results were constantly negative in liver specimens of patients without markers of HBV infection, and no reaction was observed using DNA probes lacking HBV‐DNA sequences. Detection of HBV‐DNA by in situ hybridization, using a biotinylated probe, is a rapid, reproducible, and specific histochemical method. Currently available biotinylated probes are advantageous when absolute sensitivity is not the limiting factor, and they also facilitate studies of the cellular and subcellar distribution of HBV nucleic acids.