Effect of sonication on paraffin‐embedded tissue preparation for DNA flow cytometry

Open Access
Publisher Website
Google Scholar
Abstract
Since the publication of paraffin block extraction procedures, flow cytometric analysis of DNA ploidy and S‐phase of tumor specimens has been widely applied. DNA aneuploidy, DNA tetraploid (elevated G2/M), and elevated S‐phase are clinically significant in some tumor systems. True DNA tetraploid cell lines will contain a large 4c population and perhaps an 8c population; samples with cell aggregates will also contain a 6c population. Microscopic examination of samples having a 6c peak revealed nuclei with adhering debris and doublets, triplets, and larger nuclear aggregates. After sonication, a uniform suspension of single nuclei without adherent debris was seen. In addition to reducing the percent of G2/M cells, sonication also reduced S‐phase percent such that it was closer to the bromodeoxyuridine labeling index. The DNA ploidy classification of specimens was also compared pre‐ and post‐sonication. Four of 96 breast cancer samples changed classification; all were specimens in which the histogram became cleaner and a small DNA aneuploid peak became apparent after sonication.