HBV-DNA by in situ hybridization. A method to improve sensitivity on formalin-fixed, paraffin-embedded liver biopsies

In situ hybridization of hepatitis B virus DNA by means of biotinylated probes is a rapid specific histochemical method but it is not very sensitive in formalin-fixed, paraffin-embedded material. The specificity and sensitivity of a new method using a digitonin solution and hydrolytic bath before cytohybridization in 19 formalin-fixed, paraffin-embedded liver biopsies was evaluated and a noticeable increase in staining intensivity observed. Digitonin and hydrochloric acid at low concentration increase the permeability and the diffusion of the bio-probe, improving significantly the sensitivity of reaction. Hepatocytes containing few copies of HBV-DNA were revealed by this method and microscopic visualization of specimens with numerous viral episomes was considerably improved. Therefore cytohybridization can be a specific and sensitive method, even for formalin-fixed, paraffin-embedded material, revealing active HBV replication even in HBcAg negative hepatocytes.