DNA measurement of overlapping cell nuclei in thick tissue sections

The paper describes an improved image analysis procedure for measuring the DNA content of cell nuclei in thick sections of liver tissue by absorption densitometry. Whereas previous methods only permitted the analysis of isolated nuclei, the new technique enables both isolated and overlapping nuclei to be measured. A 3D segmentation procedure determines whether each object is an isolated nucleus or a pair of overlapping nuclei; in the latter case the combined optical density is redistributed to the individual nuclei. A selection procedure ensures that only complete nuclei are measured. The method has been tested on specially-prepared Feulgen-stained 20micro sections of normal liver tissue. The overall distribution of the nuclear DNA measurements shows well-defined diploid and tetraploid peaks, with coefficient of variations of less than 10%. Similar distributions were obtained from both the isolated nuclei and overlapped nuclei sub-populations.