Non-isotopic in situ hybridisation in human pathology

Summary

Introduction

In situ hybridisation (ISH) may be defined as the direct detection of nucleic acid in intact cellular material. Nucleic acids may be exogenous or endogenous, nuclear or cytoplasmic, DNA or RNA. A variety of cell and tissue samples can be studied using ISH, from individual chromosomes in metaphase spreads to archival paraffin embedded biopsy material. Using appropriately labelled probes, the presence or absence of normal and abnormal nucleic acids can not only be detected but can also be correlated with cell and tissue morphology. This provides a wealth of information regarding both the genotype and phenotype of cells within pathological lesions and will, by combination with other techniques such as immunocytochemistry, allow greater understanding of the pathophysiology of abnormal cells and the interactions between them.

The technique of in situ hybridisation was originally described in 1969 for the detection of abundant ribosomal RNA sequences in non-mammalian systems with ³²P-labelled probes (Gall & Pardue, 1969; John et al., 1969). By increasing the sensitivity of detection and resolution of the procedure, using isotopes of high specific activity and shorter track length than ³²P (e.g. ¹²⁵I, ³⁵S, ³H), single copy genes were visualised on chromosomes (Gerhard et al., 1981). In the 1980s, non-isotopic in situ hybridisation (NISH) has been developed for gene localisation which is as sensitive as radioisotopic techniques. Single copy genes have now been mapped on chromosomes by NISH (Bhatt et al., 1988). In human disease, ISH can be applied to the detection of normal and abnormal nucleic acids. Development of techniques for ISH has been directed, in the context of laboratory medicine, towards procedures which are clinically useful.