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DNA Ploidy Investigation Aided by Image Cytometry


DNA ploidy investigation is defined here as the sequence of operations originating from paraffin embedded tumor blocks consisting of selection of tumor area, sectioning for preparation, preparation, Feulgen staining, measurement, editing, and classifying. 

DNA ploidy is a cytogenetic term describing the number of chromosome sets (n) or deviations from the normal number of chromosomes in a cell.  In cytometry, the expression is used either to describe the DNA content in a cell (c) or the total DNA distribution in a cell population.

Image cytometry is based on the Feulgen technique which is a widely used staining procedure within biology. The Schiff or Schiff-related reagent is used to bind to aldehydes created after the DNA is hydrolyzed with acid. This allows for staining of DNA in situ. The staining intensity is proportional to the DNA concentration and the amount of DNA in the nucleus is expressed by light absorbed by the Feulgen stain in the whole nucleus.

The Feulgen reaction is used to quantify the amount of DNA in a tumor nucleus. A digital camera captures images of individual nuclei in the specimen. The images are divided into image elements (picture elements - pixels). The gray tone value for each pixel represents the intensity of DNA specific staining. The value is saved in the computer which numerates between 0 (black) and 1023 (white).  The relative amount of DNA in each nucleus (DNA ploidy) is yielded by summing the optical density of all the pixels in the nucleus. DNA-ploidy measured in this way has shown to be a good prognostic and predictive method. Optical Density (OD) and Integrated Optical Density (IOD) are defined as:

During ploidy measurement, the IOD values of all of the nuclei in the specimen are registered and can be graphically developed into a histogram (x-axis: IOD value, y-axis: number of nuclei).

Clinical DNA ploidy investigations are routinely performed mainly on material from different gynecological cancers, prostate cancer, and oral cancer.


The goal of the investigation is to estimate a prognosis for the patient's cancer type. The treatment is customized depending on the ploidy result.

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