Gribko Alena, Stiefel Janis, Liebetanz Lana, Nagel Sophie Madeleine, Künzel Julian, Wandrey Madita, Hagemann Jan, Stauber Roland H, Freese Christian, Gül Désirée
Department of Otorhinolaryngology, University Medical Center Mainz, Langenbeckstr. 1, 55131 Mainz, Germany.
Fraunhofer Institute for Microengineering and Microsystems IMM, Carl-Zeiss-Str. 18-20, 55129 Mainz, Germany.
Diagnostics (Basel). 2021 Nov 4;11(11):2040. doi: 10.3390/diagnostics11112040.
detailed information about circulating tumor cells (CTCs) as an indicator of therapy response and cancer metastasis is crucial not only for basic research but also for diagnostics and therapeutic approaches. Here, we showcase a newly developed IsoMAG IMS system with an optimized protocol for fully automated immunomagnetic enrichment of CTCs, also revealing rare CTC subpopulations.
using different squamous cell carcinoma cell lines, we developed an isolation protocol exploiting highly efficient EpCAM-targeting magnetic beads for automated CTC enrichment by the IsoMAG IMS system. By FACS analysis, we analyzed white blood contamination usually preventing further downstream analysis of enriched cells.
1 µm magnetic beads with tosyl-activated hydrophobic surface properties were found to be optimal for automated CTC enrichment. More than 86.5% and 95% of spiked cancer cells were recovered from both cell culture media or human blood employing our developed protocol. In addition, contamination with white blood cells was minimized to about 1200 cells starting from 7.5 mL blood. Finally, we showed that the system is applicable for HNSCC patient samples and characterized isolated CTCs by immunostaining using a panel of tumor markers.
Herein, we demonstrate that the IsoMAG system allows the detection and isolation of CTCs from HNSCC patient blood for disease monitoring in a fully-automated process with a significant leukocyte count reduction. Future developments seek to integrate the IsoMAG IMS system into an automated microfluidic-based isolation workflow to further facilitate single CTC detection also in clinical routine.
循环肿瘤细胞（CTC）作为治疗反应和癌症转移指标的详细信息不仅对基础研究至关重要，对诊断和治疗方法也很关键。在此，我们展示了一种新开发的IsoMAG IMS系统，其具有用于CTC全自动免疫磁富集的优化方案，还能揭示罕见的CTC亚群。
我们使用不同的鳞状细胞癌细胞系，开发了一种利用高效靶向EpCAM的磁珠通过IsoMAG IMS系统进行自动CTC富集的分离方案。通过流式细胞术分析，我们分析了通常会妨碍对富集细胞进行进一步下游分析的白细胞污染情况。
发现具有甲苯磺酰活化疏水表面性质的1 µm磁珠最适合自动CTC富集。使用我们开发的方案，从细胞培养基或人血中均可回收超过86.5%和95%的加样癌细胞。此外，从7.5 mL血液开始，白细胞污染可降至约1200个细胞。最后，我们表明该系统适用于头颈部鳞状细胞癌患者样本，并使用一组肿瘤标志物通过免疫染色对分离的CTC进行了表征。
在此，我们证明IsoMAG系统能够在白细胞计数显著降低的全自动过程中从HNSCC患者血液中检测和分离CTC用于疾病监测。未来的发展旨在将IsoMAG IMS系统集成到基于微流控的自动分离工作流程中，以在临床常规中进一步促进单个CTC的检测。
