Nonradioisotopic In Situ Hybridization for HDV RNA
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The procedure described here was reported to detect the genomic strand of the hepatitis virus (HDV) RNA (
1
) in formalin-fixed, paraffin-embedded liver sections (Fig. 1 ). The method used a 27-mer end-labeled with digoxigenin (DIG). Hybrids were detected by a specific antibody coupled to alkaline phosphatase. Earlier reports had described the detection of both genomic and antigenomic HDV RNA by radioactive
35
S-labeled RNA probes (Fig. 2 ) (
2
). However, because the average number of genomic HDV RNA in each hepatocyte is very high (around 300,000 copies/cell) (
3
), HDV is the ideal target to be detected by a nonradioactive
in situ
hybridization procedure, even using a short oligonucleotide probe, i.e., representing only approx 1.6% of the total length of the genome.
Fig. 1.
Detection of genomic HDVRNA in nuclei of hepatocytes by nonradioisotopic
in situ
hybridization using a digoxigenin-labeled synthetic oligonucleotide. Counterstaining with methyl green. Original magnification �40.
Fig. 2.
Detection of genomic HDV RNA in nuclei of hepatocytes by
in situ
hybridization using
35
S-labeled RNA probes. Counterstaining with hematoxylin. Original magnification �100.
