Wang Xuebin, Sun Liying, Zhang Haiming, Wei Lin, Qu Wei, Zeng Zhigui, Liu Ying, Zhu Zhijun
Department of General Surgery, Beijing Friendship Hospital, Capital Medical University, Beijing, P. R. China.
Liver Transplantation Center, National Clinical Research Center for Digestive Diseases (NCRC-DD), Beijing Friendship Hospital, Capital Medical University, Beijing, P. R. China.
PeerJ. 2019 Apr 1;7:e6681. doi: 10.7717/peerj.6681. eCollection 2019.
We aimed to generate a capture platform that integrates a deterministic lateral displacement (DLD) microfluidic structure with magnetic-activated cell sorting (MACS) technology for miniaturized, efficient, tumor antigen-independent circulating tumor cell (CTC) separation.
The microfluidic structure was based on the theory of DLD and was designed to remove most red blood cells and platelets. Whole Blood CD45 MicroBeads and a MACS separator were then used to remove bead-labeled white blood cells. We established HepG2 human liver cancer cells overexpressing green fluorescent protein by lentiviral transfection to simulate CTCs in blood, and these cells were then used to determine the CTC isolation efficiency of the device. The performance and clinical value of our platform were evaluated by comparison with the Abnova CytoQuest™ CR system in the separating of blood samples from 12 hepatocellular carcinoma patients undergoing liver transplantation in a clinical follow-up experiment. The isolated cells were stained and analyzed by confocal laser scanning microscopy.
Using our integrated platform at the optimal flow rates for the specimen (60 µl/min) and buffer (100 µl/min per chip), we achieved an CTC yield of 85.1% ± 3.2%. In our follow-up of metastatic patients, CTCs that underwent epithelial-mesenchymal transition were found. These CTCs were missed by the CytoQuest™ CR bulk sorting approach, whereas our platform displayed increased sensitivity to EpCAM CTCs.
Our platform, which integrates microfluidic and MACS technology, is an attractive method for high-efficiency CTC isolation regardless of surface epitopes.
我们旨在构建一个捕获平台，该平台将确定性侧向位移（DLD）微流控结构与磁激活细胞分选（MACS）技术相结合，以实现小型化、高效、不依赖肿瘤抗原的循环肿瘤细胞（CTC）分离。
微流控结构基于DLD理论设计，旨在去除大多数红细胞和血小板。然后使用全血CD45微珠和MACS分离器去除珠标记的白细胞。我们通过慢病毒转染建立了过表达绿色荧光蛋白的HepG2人肝癌细胞，以模拟血液中的CTC，然后使用这些细胞来确定该装置的CTC分离效率。在一项临床随访实验中，通过与Abnova CytoQuest™ CR系统比较，评估了我们平台在分离12例接受肝移植的肝细胞癌患者血样中的性能和临床价值。对分离出的细胞进行染色，并通过共聚焦激光扫描显微镜进行分析。
使用我们的集成平台，在样本的最佳流速（60 μl/min）和缓冲液（每芯片100 μl/min）下，我们实现了85.1%±3.2%的CTC回收率。在对转移性患者的随访中，发现了经历上皮-间质转化的CTC。这些CTC被CytoQuest™ CR批量分选方法遗漏了，而我们的平台对EpCAM CTC显示出更高的敏感性。
我们的平台集成了微流控和MACS技术，是一种无论表面表位如何都能高效分离CTC的有吸引力的方法。
