| Objective: Explore to develop a method for detecting circulating tumor cells (CTCs) in peripheral blood of stage I lung cancer patients with high sensitivity and specificity, which may benefit for detecting micrometastasis in early stage, assessment of prognosis, monitoring relapse and metastasis, selecting appropriate treatment and judging therapeutic effect.Methods: 35 cases of initial treatment patient who underwent resection and diagnosed to be non-small cell lung cancer (NSCLC) by biopsy were studied, including 15 patients at stage I, 9 patients at stage II, 9 patients at stage III, 2 patients at stage IV, 22 patients of squamous carcinoma, 13 patients of adenocarcinoma. As a control group, 20 cases of benign pulmonary disease and 20 cases of healthy volunteers were selected. In order to detect circulating tumor cells of peripheral blood in patients with early stage NSCLC, immunomagnetic separation (IMS) was performed on peripheral blood specimens of early stage patients (stage I), advanced stage patients (stage II ~ FV), benign pulmonary disease patients and healthy volunteers respectively, using nanometer immunomagnetic beads coated with anti-CK7/8 (cytokeratin, CK) monoclonal antibody. Then immunocytochemistry (ICC) was performed on immunostain tumor cells or abnormal epithelial cells enriched by IMSusing anti-pan-CK antibody (AE1/AE3). At the same time, the detection of CK positive cell using IMS followed by ICC was compared with the detection of CK positive cell using simple ICC and the detection of CK19mRNA or CEAmRNA using nest RT-PCR method. We also followed up postoperative NSCLC patients in early stage (stage I) and evaluated the correlation of detecting CTCs and clinical prognosis hi patients with NSCLC.Results: 1. The detection rate of CK positive cells using IMS followed by ICC method was 33.3% (5/15), the detection rate of CK positive cells using simple ICC method was 0% (0/15) and the detection rate of CK19mRNA and CEAmRNA using nest RT-PCR method was 46.7% (7/15) and 40.0% (6/15) respectively in the specimens of patients with stage I lung cancer. 2. The detection rate of CK positive cells using IMS followed by ICC method was 50% (10/20), the detection rate of CK positive cells using simple ICC method was 0% (0 / 20) , the detection rate of CK19mRNA and CEAmRNA using nest RT-PCR method was 65% (13/20) and 60% (12/20) respectively in the specimens of patients with advanced stage lung cancer. 3. The detection rate of CK positive cells using IMS followed by ICC method and simple ICC method was the same 0%(0/20), the detection rate of CK19mRNA and CEAmRNA using nest RT-PCR method was 40% (8/20) and 30%(6/20) respectively in the specimens of patients with benign pulmonary disease. 4. The detection rate of CK positive cells using IMS followed by ICC method and simple ICC method was the same, 0%(0/20), the detection rate of CK19mRNA and CEAmRNA using nest RT-PCR method was 30%(6/20) and 25%(5/20) respectively in the specimens of healthy volunteers. 5. 5 patients were positive for CK cells preoperative using IMS followed by ICC methodwere detected for CK cells again one month after operation in the patients with stage I lung cancer. 2 of the 5 cases were still positive, the other 3 cases were negative. In the next 6 - 9 month follow-up, 10 patients that were negative for CK cells before operation and 3 patients that were positive for CK cells before operation but negative after operation were all in good condition, whereas 2 patients that were positive for CK cells before and after operation all suffered from relapse and metastasis.Conclusion: IMS is a sensitive method to capture CTCs of lung cancer. The detection of CTCs suggests that it may be the existence of micrometastasis in early stage lung cancer, and the detection of CTCs in lung cancer patients is closely related to the clinical prognosis of patients. Our results revealed that detection of CTCs hi lung cancer patients using IMS technique has important value to prognostic assessment, discovery of early stage microm...
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