Establishment Of The Method For Micrometastasis Of CEA Producing Tumors And Its Clinical Significance

In our study, we try to establish a method for detecting micrometastases of CEA-producing tumors and evaluate its clinical significance. First, nucleated cells were separated from fresh peripheral blood and the total RNA was isolated from nucleated cells. Reverse transcriptions were performed with the total RNA as template using Primers (T) and (2) respectively to produce cDNA. Then PCR was carried out further with cDNA as template to obtain the given CEA DNA fragment. The resultants of RT-PCR were analyzed on 5% polyacrylamide gel electrophoresis and by restriction analysis. Finally they were sequenced in order to verify the reliability. Results: OD 260/280 values of the total RNA from nucleated cells in peripheral blood were between 1.5-1.8. Intact 28S and 18S RNAs of the total RNA were seen clearly on gel electrophoresis. The positive samples of RT-PCR products showed a band in 131bp and 338bp respectively on electrophoresis. The former was turned to 59bp and 72bp fragments by Mva I digestion. The results of DNA sequencing completely coincide with the sequence of CEA cDNA in literature. 25 of 46 patients with CEA-producing tumors were positive for CEA mRNA by RT-PCR. 1 of 5 non-CEA-producing tumors showed positive. None of 12 healthy volunteers had positive results. The presence of CEA mRNA in blood was related to tumor distant metastases(/?<0.005). After a half year's follow-up, distant metastases occurred in 3 patients without distant metastases who had positive CEA mRNA. So detecting CEA mRNA by RT-PCR may predict early metastases of tumor cells. The CEA mRNA positive rate increased with tumor clinical staging(p<0.005). There were no significant differences indicated between ages, serum CEA and CEA mRNA(p>0.05). 2 patients with negative CEA mRNA preoperatively were detected positive postoperatively, which indicates that operation may encourages tumor cells to shed into the blood stream.