目的

本研究旨在建立一种高效的结直肠癌循环肿瘤细胞（CTC）分选系统，并探讨CTC代替肿瘤组织进行基因突变检测的可行性。

方法

建立EpCAM与Vimentin抗体修饰的CTC分选系统。通过细胞实验检测CTC分选系统对结直肠癌细胞的分选效率，并对30例结直肠癌患者的CTC进行捕获计数。利用NGS检测患者肿瘤组织样本中热点突变基因，并通过Sanger测序检测血液CTC中热点突变基因，分析肿瘤组织与血液CTC热点基因突变的一致性。

结果

本研究构建的CTC分选系统具有较好的分散性、稳定性及较低的细胞毒性，特异度为90.54%，敏感度为89.07%。结直肠癌患者血液中CTC平均数为（10.90±5.11）个/7.5 mL。通过CTC检出的高频突变基因（TP53、KRAS、EGFR、BRAF、APC、PIK3CA）与组织检测均具有较高的一致性，所有基因总体突变的一致率为（92.89±4.89）%。

结论

基于Ep-LMB/Vi-LMB依次富集的CTC分选系统能够对结直肠癌患者外周血中CTC进行高效捕获。将捕获的CTC用于临床上的热点基因突变检测，在一定程度上解决了肿瘤组织取材困难的问题，具有重要的临床意义。

Objective

The aim of this study was to establish an efficient circulating tumor cells（CTC） sorting system for colorectal cancer and to investigate the feasibility of CTC instead of tumor tissue for gene mutation detection.

Methods

The CTC sorting system modified by EpCAM and Vimentin antibody was established. The sorting efficiency of the CTC sorting system on colorectal cancer cells was detected by cellular experiments， and CTC from 30 colorectal cancer patients were captured and counted. The hotspot mutated genes in patient tumor tissue samples were detected by NGS， and the hotspot mutated genes in blood CTC were detected by Sanger sequencing， and the consistency of hotspot gene mutations between tumor tissues and blood CTC was analyzed.

Results

The CTC sorting system constructed in this study had good dispersion， stability and low cytotoxicity， with a specificity of 90.54% and a sensitivity of 89.07%.The mean number of CTC in the blood of colorectal cancer patients was （10.90±5.11）/7.5 mL. The highfrequency mutated genes （TP53， KRAS， EGFR， BRAF， APC， PIK3CA） detected by CTC were all in high concordance with the tissue assay， and the overall mutation concordance rate for all the genes was （92.89±4.89）%.

Conclusions

The CTC sorting system based on sequential enrichment of Ep-LMB/Vi-LMB is capable of efficiently capturing CTC from peripheral blood of colorectal cancer patients. Using the captured CTC for hotspot mutation detection in the clinic solves the problem of difficult tumor tissue sampling to a certain extent， which is of great clinical significance.