The effects of formalin fixation on the detection of apoptosis in human brain by in situ end-labelling of DNA

Abstract

The technique of DNA in situ end-labelling (ISEL) for the detection of apoptotic cells has recently become the method of choice. The incorporation of a labelled nucleotide to facilitate detection into the single-stranded region of DNA cleaved by endogenous nucleases has proved to be a sensitive and straightforward technique. Previous reports have applied the technique to the study of apoptotic cells in brain tissue, which is normally subjected to relatively long-term formalin fixation. In this study we have examined the effects of long-term formalin fixation on the ability to detect apoptosis using ISEL in a variety of pathologies and in a normal rat testis. In the tissues which had been treated with overnight formalin fixation, apoptotic cells were readily identified in those pathologies where it might be expected to occur. However, in tissue which had been fixed for several weeks or more, apoptotic cells were not detectable. Samples of brain lymphoma tissue and rat testis subjected to a prospective analysis with respect to fixation time showed that the ability to detect apoptotic cells tailed off at around 3-5 weeks. In order to obviate the risk of false negative results it would be desirable to use ISEL in tissues formalin fixed for less than this period.