Isolation and characterization of circulating tumor cells using a novel workflow combining the CellSearch® system and the CellCelector™ |
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| Authors: | Martin Horst Dieter Neumann Helen Schneck Yvonne Decker Susanne Schömer André Franken Volker Endris Nicole Pfarr Wilko Weichert Dieter Niederacher Tanja Fehm Hans Neubauer |
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| Affiliation: | 1. Department of Obstetrics and Gynecology, University Hospital and Medical Faculty of the Heinrich‐Heine University, Duesseldorf, Germany;2. Institute of Pathology, University Hospital Heidelberg, Heidelberg, Germany;3. Institute of Pathology, Technical University Munich, Munich, Germany;4. National Center for Tumor Diseases (NCT), Heidelberg, Germany;5. Member of the German Cancer Consortium |
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| Abstract: | Circulating tumor cells (CTC) are rare cells which have left the primary tumor to enter the blood stream. Although only a small CTC subgroup is capable of extravasating, the presence of CTCs is associated with an increased risk of metastasis and a shorter overall survival. Understanding the heterogeneous CTC biology will optimize treatment decisions and will thereby improve patient outcome. For this, robust workflows for detection and isolation of CTCs are urgently required. Here, we present a workflow to characterize CTCs by combining the advantages of both the CellSearch® and the CellCelector? micromanipulation system. CTCs were isolated from CellSearch® cartridges using the CellCelector? system and were deposited into PCR tubes for subsequent molecular analysis (whole genome amplification (WGA) and massive parallel multigene sequencing). By a CellCelector? screen we reidentified 97% of CellSearch® SKBR‐3 cells. Furthermore, we isolated 97% of CellSearch®‐proven patient CTCs using the CellCelector? system. Therein, we found an almost perfect correlation of R2 = 0.98 (Spearman's rho correlation, n = 20, p < 0.00001) between the CellSearch® CTC count (n = 271) and the CellCelector? detected CTCs (n = 252). Isolated CTCs were analyzed by WGA and massive parallel multigene sequencing. In total, single nucleotide polymorphisms (SNPs) could be detected in 50 genes in seven CTCs, 12 MCF‐7, and 3 T47D cells, respectively. Taken together, CTC quantification via the CellCelector? system ensures a comprehensive detection of CTCs preidentified by the CellSearch® system. Moreover, the isolation of CTCs after CellSearch® using the CellCelector? system guarantees for CTC enrichment without any contaminants enabling subsequent high throughput genomic analyses on single cell level. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:125–132, 2017 |
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| Keywords: | circulating tumor cells CellCelector™ CellSearch® single cell analysis |
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