DNA Ploidy 
Contents
Ploidy is a cytogenetic term used to describe the number of chromosome sets, or deviation from the normal number of chromosomes in a cell. DNA ploidy is another objective expression for large scale genomic instability which discriminates between tumour samples which have normal, stable DNA amounts and samples with abnormal and usually unstable DNA amounts.
Overview
DNA ploidy analysis is a method where the total DNA content per nuclei for all the nuclei in a tumour sample is calculated. The DNA ploidy analysis has traditionally been done by flow cytometry. We have developed an image analysis system for DNA ploidy analyses based on the Feulgen technique, which is a widely used staining method in biology. The Feulgen reaction is used to quantify the DNA ploidy distribution in tumor nuclei. DNA ploidy is used in diagnostics in gynecological cancers and is a promising prognostic marker in several other cancer types.
DNA ploidy in cell cycle
Normal, somatic cells have in their G0/G1 phase a complete set of chromosomes (diploid condition). This set consists of 23 chromosomes (haploid, n) times two that is a total of 46 chromosomes (diploid, 2n). If we were to run a DNA ploidy analysis on normal, somatic cell nuclei, we would find the majority of nuclei in the G0/G1 phase. At the same time, some of the nuclei are always undergoing cell division and we would find nuclei in the G2 phase, having a double set of chromosomes (tetraploid, 4n). We would probably also see some nuclei undergoing the replication phase (S-phase). When we plot these nuclei in a histogram it would look something like this:
Automatic classification of cell types
A condition for automatic DNA ploidy analysis is that one must classify nuclei in relation to the cell type it was isolated from (epithelial nuclei or nuclei from connective tissue, as well as lymphocytes and plasma cells from the tumour’s blood supply). A drastic benefit with a system based on scanner technology, is that one can relatively quickly measure all of the nuclei in the specimen (>10,000). The manual editing job is however time-consuming, and we have therefore developed a classification algorithm which classifies the different nuclei automatically with a correct classification rate between 80 and 90%. The remaining nuclei are edited manually.
Automatic classification of DNA histograms
DNA histograms are traditionally classified manually. As the last phase in automation, we have developed an algorithm for automatic classification of such histograms.
Prognostic markers
DNA Ploidy material preperation process
Several studies have been done on DNA ploidy in cancer cells and tumours. It is well known that aneuploidy is a sign of malignancy, and often associated with a poor prognosis for the patient. We have shown that DNA ploidy, in form of non-diploid conditions, can be correlated with a poorer clinical outcome for patients with gynecological, prostate and colorectal cancers, and patients with Barrett’s esophagus.
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