IMPROVED METHOD FOR RELEASE OF CELL-NUCLEI FROM PARAFFIN-EMBEDDED CELL MATERIAL OF SQUAMOUS-CELL CARCINOMAS

An improved method of releasing cell nuclei from paraffin-embedded hig hly keratinized squamous cell carcinomas by pretreatment with 85% form ic acid-0.3% H2O2 followed by enzymatic treatment with subtilisin Carl sberg is described, After DAPI staining the resulting suspensions of c ell nuclei were analyzed by DNA flow cytometry, in addition to microsc opy. The total yield of released cell nuclei was improved and the prop ortion of tumor cell nuclei in the suspensions increased from 12% to 5 3% using this method compared to the conventional preparation techniqu e. Cytoplasmic residue disappeared nearly completely. Thus, the findin g of false aneuploid cell populations representing diploid cells with autofluorescent cytoplasm could be avoided. In addition, even small tr ue aneuploid cell populations could be detected due to the increased p roportion of released tumor cell nuclei and the lower proportion of ba ckground in the >2C region. (C) 1993 Wiley-Liss, Inc.