Application of silver acetate autometallography and gold-silver staining methods for in situ DNA hybridization

In situ hybridization using biotinylated DNA probes has become an important tool in histopathology. It is well known that the sensitivity of the methods used to demonstrate viral DNA in formalin-fixed and paraffin-embedded specimen depends strongly on the detection system used. In the present study, an optimized in situ DNA hybridization protocol was combined with four different approaches of gold-silver staining methods. For silver enhancement, the recently described method of silver acetate autometallography, a technique allowing highly efficient development without the necessity of dark room illumination has been used. The most efficient detection method found in our experiments was the use of gold-adsorbed anti-biotin antibodies with subsequent silver enhancement. This staining procedure can be completed in 5 hours including hybridization and is a highly sensitive alternative to peroxidase and alkaline phosphatase detection systems.