Cytofluorometric nuclear DNA content analysis of breast tissues after frozen section diagnosis

OBJECTIVE: To establish a suitable method for measurement of nuclear DNA co ntent in breast tissues from frozen storage after frozen section diagnosis. STUDY DESIGN: For fundamental research, rat liver samples preserved ill a d eep freezer were used. Four protocols were used (1. fixation with 70% ethan ol followed by naked nuclei preparation; 2. fixation with 10% neutral buffe red formalin followed by naked nuclei preparation; 3. preparation for linke d nuclei prior to fixation with 70% ethanol; and 4. preparation for naked n uclei prior to fixation with 70% neutral buffered formalin). For clinical r esearch, 13 separate fresh frozen breast tissue samples were analyzed after frozen section diagnosis. One contained a malignant phyllodes tumor (MPT) consisting of 2 components, benign epithelial cells and malignant stromal c ells; 3 were benign tumors containing fibroadenoma; ann 9 cases were carcin omas, consisting of 5 scirrhous, 3 papillotubular and 1 mucinous. RESULTS: Protocols 1, 2 and 3 were not suitable methods for our purpose bec ause remaining cytoplasm or cohesive nuclei were observed. Ill protocol 4 t he cytoplasm was completely undetectable, and nuclei were suitably separate d for nuclear DNA content measurement. Benign epithelial cell component nuc lei presented a diploid pattern, and the malignant stromal cell component n uclei indicated a euploid pattern in MPT. All 3 cases of benign constituent s in fibroadenoma showed a diploid pattern, as did the 3 carcinoma cases (1 mucinous, 1 scirrhous and 1 papillary). Four scirrhous and 2 papillary car cinomas showed an aneuploid pattern. CONCLUSION: Our findings show that it is possible to measure nuclear DNA co ntent of human frozen storage tissues after frozen section diagnosis.