中国现代医学杂志2017，Vol.27Issue(19)：34-39,6.DOI:10.3969/j.issn.1005-8982.2017.19.007
外周血捕获循环肿瘤细胞的有效方法研究
Isolation of single circulating tumor cell from peripheral blood of liver cancer patients by immunomagnetic beads and micromanipulation
摘要
Abstract
Objective To isolate single circulating tumor cell (CTC) from peripheral blood of liver cancer patients.Methods Four commercial kits including OncoQuick,Ficoll-Paque PLUS (Ficoll),RosetteSep and CELLection,as well as Ficoll-CELLection-combination were used to enrich DiI-stained SW480 cells added to peripheral blood of healthy adults.In order to compare the above five methods,recovery and purity of enriched tumor cells were counted or observed.Then,detection cells including NCI-H460,HCT-116,DLD-1,SW480 and TF-1,and CTCs from the peripheral blood of liver cancer patients were isolated by micromanipulation individually.Subsequently,mRNA expression level of CD45 antigen,the leukocyte common antigen,was detected by qRT-PCR.Results Compared in pairs,the recovery of the five methods [OncoQuick (52.5 ± 3.5)％,Ficoll (46.6 ± 1.9)％,RosetteSep (36.9 ± 5.5)％,CELLection (14.1-± 3.1)％ and Ficoll-CELLection-combination (8.8-± 1.4)％] revealed significant differences (P ＜ 0.05) except for OncoQuick and Ficoll (P ＞ 0.05),CELLection and Ficoll-CELLection-combination (P ＞ 0.05) by one-way ANOVA test and SNK-q method.Moreover,CELLection and Ficoll-CELLection-combination enriched tumor cells with higher purity when they were observed under fluorescent microscope.In addition,CD45 mRNA expression of all single detection cells and CTC were negative except TF-1 (mean Ct within 60 cycles:TF-1 (34.21 ± 0.22),GAPDH (29.94 ± 0.59),others none;n =3).Conclusions The combination of immunomagnetic beads and micromanipulation is an effective method to isolate single CTC from peripheral blood of liver cancer patients.The captured single CTC keeps its original molecular characteristics for subsequent single-cell analysis.Furthermore,our method lays a foundation for clinical application of CTC,for tumor molecular characteristic analysis,therapeutic effect evaluation and treatment monitoring.关键词
循环肿瘤细胞/免疫磁珠法/显微操作Key words
circulating tumor cell/immunomagnetic beads/micromanipulation分类
医药卫生引用本文复制引用
周洲,李良平,高采平,钟应佳,翟祎星,赖兵..外周血捕获循环肿瘤细胞的有效方法研究[J].中国现代医学杂志,2017,27(19):34-39,6.基金项目
四川省科技厅资助项目(No:2014TD0028) （No:2014TD0028）
