Background: The prognostic value of flow cytometric DNA analysis on paraffin-embedded tumor samples has been controversial in esophageal cancer. To clarify its true significance, the authors developed an improved method that excludes the possibility of contamination by lymphocytes in tumor sample.
Methods: Single nuclear suspension was prepared from paraffin-embedded samples on 103 patients with squamous cell carcinoma of the esophagus. Both DNA content and nuclear size were simultaneously measured by flow cytometry on 30,000 nuclei, and contaminated lymphocyte nuclei were eliminated from the data by optimal gating. Correlation between DNA ploidy and postoperative survival was examined.
Results: Analysis using a flow cytometric cell sorter showed that the frequency of tumor cells in the lymphocyte-reducing gating fraction (LGF) was significantly higher than that in the conventional nongating fraction (NGF). LGF analysis showed aneuploid peaks in 58 patients (56.3%), but NGF analysis showed aneuploid peaks in only 38 patients. LGF analysis revealed that the aneuploid tumors had higher histologic grading (P < 0.05) and worse survival rate (P < 0.01) compared with diploid tumors. However, conventional methods could not detect this difference.
Conclusions: Flow cytometric analysis gating by nuclear size may be helpful to detect aneuploid peaks, and for predicting prognosis of patients with squamous cell carcinoma of esophagus.