Division of Hematology/Oncology and Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, CA, United States.
Methods. 2013 Dec 1;64(2):114-8. doi: 10.1016/j.ymeth.2013.07.029. Epub 2013 Jul 26.
Circulating tumor cells (CTCs) are cells shed by the primary tumor into the blood stream capable of initiating distant metastasis. In the past decade, numerous assays have been developed to reliably detect these extremely rare cells. However, methods for purification of CTCs with little or no contamination of normal blood cells for molecular profiling are limited. We have developed a novel protocol to isolate CTCs by combining immunomagnetic enrichment and fluorescence-activated cell sorting (IE/FACS). The two-part assay includes (1) immunomagnetic capture using magnetic beads conjugated to monoclonal antibody against an epithelial cell adhesion marker (EpCAM) to enrich for tumor cells; and (2) FACS analysis using EpCAM to purify tumor cells away from mononuclear cells of hematopoietic lineage. Downstream molecular analyses of single and pooled cells confirmed the isolation of highly pure CTCs with characteristics typical that of malignant cells.
循环肿瘤细胞（CTCs）是原发性肿瘤脱落到血液中能够引发远处转移的细胞。在过去的十年中，已经开发出许多可靠地检测这些极其罕见细胞的方法。然而，用于纯化 CTCs 的方法很少或没有污染正常血细胞进行分子分析是有限的。我们开发了一种新的方案，通过结合免疫磁珠富集和荧光激活细胞分选（IE/FACS）来分离 CTCs。两步法包括（1）使用与针对上皮细胞黏附标志物（EpCAM）的单克隆抗体偶联的磁珠进行免疫磁珠捕获，以富集肿瘤细胞；和（2）使用 EpCAM 通过 FACS 分析从造血谱系的单核细胞中纯化肿瘤细胞。对单个和汇集细胞的下游分子分析证实了高度纯净的 CTCs 的分离，其特征与恶性细胞典型特征一致。
