Flow cytometric immunophenotyping of serous effusions and peritoneal washings: comparison with immunocytochemistry and morphological findings

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Abstract
Aim—To evaluate immunophenotyping by means of flow cytometry as a complementary method for the detection of malignant cells in serous effusions and peritoneal washings. Material and methods—Frozen samples of 49 fresh serous effusions and peritoneal washings were analysed by flow cytometry, using monoclonal antibodies against CD45, Ber-EP4, and N-cadherin. Results were compared with smear and cell block morphology, as well as immunocytochemistry on paraffin wax embedded cell blocks. Results—Seventeen specimens were cytologically diagnosed as malignant, whereas 25 were interpreted as benign. The remaining seven specimens were diagnosed as indeterminate or suspicious for malignancy. Ber-EP4 positive cells were detected in 16 of the 17 cytologically malignant effusions, as well as in five of seven suspicious cases and five of 25 specimens with benign cytology. In the latter group, three specimens showed atypical or malignant cell groups that were missed in routine morphological evaluation. In two additional samples, obtained from patients with benign and borderline ovarian tumours, Ber-EP4 positive cells showed benign or mildly atypical features, and were interpreted as exfoliated benign or borderline malignant epithelial cells of tubal origin, or as endosalpingiosis. All five Ber-EP4 positive indeterminate specimens showed atypical or malignant cells on re-evaluation, and were Ber-EP4 positive in four of five cases using immunohistochemistry in cell block sections. Large numbers of CD45 positive and relatively few N-cadherin positive cells were detected in most specimens with the use of flow cytometry, when compared with morphological evaluation. Conclusions—Flow cytometry is a rapid and highly effective method for the evaluation of effusions and peritoneal washings. The detection of Ber-EP4 positive cells using flow cytometry is strongly indicative of the presence of carcinoma cells in effusions and peritoneal washings. Although false positives are relatively infrequent, all specimens should be carefully evaluated morphologically to prevent the diagnosis of benign epithelial clusters as malignant.