An improved hedley method for preparation of paraffin-embedded tissues for flow cytometric analysis of ploidy and S-phase

✍ Scribed by Thomas Heiden; Naining Wang; Dr. Bernhard Tribukait

• Publisher: John Wiley and Sons
• Year: 1991
• Tongue: English
• Weight: 616 KB
• Volume: 12
• Category: Article
• ISSN: 0196-4763
• DOI: 10.1002/cyto.990120705

✦ Synopsis

A modification of the Hedley-method for flow cytometric DNA analysis of paraffin-embedded tissues is presented. Dewaxed and dehydrated tissue from paraffin blocks was incubated with subtilisin Carlsberg (pronase, Sigma protease XXIV) and then stained directly without washing and centrifugation. The loss of material was minimized, which was advantageous, for example, for the analysis of core-biopsies, and all measured samples showed extremely low frequencies of clumped cell nuclei. This made it eas-ier to detect polyploid nuclei and even rare nuclei of high ploidy could be identified. S-phase analyses were more precise, since the background originating from clumped debris particles was very low. The improved method was applied to the estimation of frequencies of highpolyploid nuclei found in various diploid, tetraploid, and aneuploid human myosarcomas of the uterus.