Background
Free disseminated peritoneal tumor cells derive from the detachment from primary cancer and may result in peritoneal carcinomatosis. Peritoneal lavage cytology has low sensitivity in detecting free peritoneal tumor cells; reverse-transcriptase polymerase reaction showed higher sensitivity but low specificity. Our study introduces the combination of the RT-PCR, the immunomagnetic enrichment and the immunofluorescence for the detection of peritoneal free tumor cells.
Materials and Methods
Samples of peritoneal lavage were collected from 22 gastric and 45 colorectal cancer patients; samples were also obtained from 6 patients who underwent abdominal surgery for non-malignant diseases. CEA and CK20 mRNA levels were quantified using a real-time qRT-PCR system. Immunomagnetic enrichment followed by immunofluorescence analysis was performed using monoclonal antibody against the pan-epithelial marker EpCAM/CD326 and polyclonal antibodies against the carcinoembryonic antigen.
Results
For gastric carcinoma the positivity rate for cytology, immunofluorescence and qRT-PCR was 14%, 18% and 77% respectively; for colorectal carcinoma the positivity rate for cytology, immunofluorescence and qRT-PCR was 0%, 18% and 44% respectively. All patients except one when positive at immunofluorescence were also positive at qRT-PCR. All samples of peritoneal lavage from the control group resulted negative for cytology, IF and real time qRT-PCR.
Conclusion
The combination of conventional real time qRT-PCR with immunoenrichment and immunofluorescence, which permit morphological assessment and unequivocal identification of the DTCs as well as validation of the molecular analysis, could be an useful and more powerful procedure for the detection of free peritoneal tumor cells
