In-Situ End Labelling with Bromodeoxyuridine - an Advanced Technique for the Visualization of Apoptotic Cells in Histological Specimens

Abstract

The fragmentation of nuclear DNA is considered to be a biochemical indication of programmed cell death (PCD) in various tissues. Several light microscopical approaches have been employed for the visualization of such fragmentation and have used either in-situ nick end labelling (ISEL) or in-situ nick translation (ISNT) whith labelled dUTP. In the current paper, we can report a new method in which 0.5 µm Epon sections of mouse jejunum were incubated with terminal deoxynucleotidyl transferase (TdT) and 5-bromo-2-deoxyuridine (BrdU) for the in-situ tailing of fragmented DNA. Anti BrdU and the peroxidase anti peroxidase method (PAP) were used for staining the reaction product. Semithin sections of jejunum revealed high resolution images of labelled nuclei in the intestinal mucosa. An examination with confocal laser scanning microscopy indicated the differential distribution of immunoprecipitates in cellular nuclei, perhaps representing those distinct levels of DNA cleavage that may characterise the discrete stages of apoptosis. This new method is likely to offer new possibilities for the high resolution imaging of PCD, including the assessment of time courses and apoptosis statuses in histochemically characterized cells.

Key words

In-situ Tailing - Semithin Section - Bromodesoxyuridine - Apoptosis - DNA Fragmentation