Department of Microsystems Engineering, University of Freiburg , Georges-Köhler-Allee 103, D-79110 Freiburg, Germany.
GILUPI GmbH , Hermannswerder 20a, 14473 Potsdam, Germany.
Anal Chem. 2017 Feb 7;89(3):1846-1854. doi: 10.1021/acs.analchem.6b04219. Epub 2017 Jan 12.
The detection of circulating tumor cells (CTCs) in the blood of cancer patients is a challenging task. CTCs are, especially at the early stages of cancer development, extremely rare cells hidden in a vast background of regular blood cells. We describe a new strategy for the isolation of CTCs from whole blood. The key component is a medical wire coated with a multilayer assembly that allows highly specific capture of EpCAM (epithelial cell adhesion molecule) positive CTCs from blood. The assembly is generated in a layer-by-layer fashion through photochemically induced C,H insertion reactions and consists of a protective layer, which shields the contacting solution from the metal, a protein resistant layer, which prevents nonspecific interactions with proteins and a layer containing the EpCAM antibodies. In vitro experiments show that these surfaces can capture tumor cells from whole blood with enrichment factors (specifically vs nonspecifically bound cells) of up to about 3000 compared to the number of leucocytes in the blood. The purity of the isolated cells is greater than 90%. After "fishing" them from the blood, the cells, still bound to the wire, can be genetically analyzed. This demonstrates that this strategy might prove useful for next generation sequencing.
从癌症患者血液中检测循环肿瘤细胞 (CTC) 是一项具有挑战性的任务。CTC 是在癌症发展的早期阶段极其罕见的细胞，隐藏在大量的正常血细胞背景中。我们描述了一种从全血中分离 CTC 的新策略。关键组件是一种涂有多层组件的医用金属丝，该组件允许通过光化学诱导的 C,H 插入反应高度特异性地捕获 EpCAM（上皮细胞黏附分子）阳性的 CTC。该组件通过逐层方式生成，包括一个保护层，该保护层将接触溶液与金属隔开；一个蛋白质抗性层，该层防止与蛋白质发生非特异性相互作用；以及一个包含 EpCAM 抗体的层。体外实验表明，与血液中的白细胞数量相比，这些表面可以从全血中捕获肿瘤细胞，其富集因子（特异性结合细胞与非特异性结合细胞之比）高达约 3000。分离出的细胞纯度大于 90%。从血液中“捕获”这些细胞后，仍然与金属丝结合的细胞可以进行基因分析。这表明该策略可能对下一代测序有用。
