The demonstration of viral DNA in human tissues by in situ DNA hybridisation

Summary

Introduction

The extraction of single-stranded human DNA, its separation by gel electrophoresis, its transfer to a nitrocellulose filter (the Southern blot) followed by hybridisation with a radioactively labelled viral probe consisting of a complementary DNA sequence forms the fundamental basis of the now well-established technique of DNA hybridisation. Dot blot hybridisation is a, more rapid variation of this basic technique.

However, the histopathologist is concerned with morphology and topographical relationships within tissue sections and the investigation of DNA bands does not allow one to precisely localise viral DNA within human tissues. But now the methods of molecular biology and immunohistochemistry have come together to allow the specific recognition of viral DNA in tissue sections.

Since 1985 there has been an increasing number of publications on the subject of in situ hybridisation for human papillomavirus (HPV) DNA. Our own work in this field was always aimed at developing a technique that was applicable to the routinely fixed and processed material in a hospital diagnostic histopathology laboratory (Lewis et al., 1987; Wells et al., 1987). Such a technique would permit the retrospective evaluation of archival material, thus considerably widening the scope for investigating the viral status of a variety of pathological lesions.

Numerous detection systems have been developed to visualise labelled gene probes. Autoradiography has been used to detect radio-labelled probes in cell smears and paraffin sections (Gupta et al., 1985). Although autoradiographical techniques are very sensitive, the use of radio-labelled material is unsuitable for most routine applications.