An integrated microfluidic chip system for single-cell secretion profiling of rare circulating tumor cells.
An integrated microfluidic chip system for single-cell secretion profiling of rare circulating tumor cells.
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用于稀有循环肿瘤细胞单细胞分泌分析的集成微流控芯片系统
DOI：
10.1038/srep07499
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发表时间：
2014-12-16
影响因子：
4.6
通讯作者：
Shi Q
中科院分区：
文献类型：
--
作者：
Deng Y;Zhang Y;Sun S;Wang Z;Wang M;Yu B;Czajkowsky DM;Liu B;Li Y;Wei W;Shi Q
已经证明了单个循环肿瘤细胞（CTC）的遗传和转录谱分析以及表面标志物鉴定。然而，由于分离的CTC群体的有限纯度和缺乏处理和分析稀有CTC的单细胞蛋白质组学方法，尚未实现在单个CTC分辨率下的功能蛋白质的定量分析。在这里，我们开发了一种集成的微流体系统，专门用于简化全血中CTC的分离、纯化和单细胞分泌组学分析。该平台的关键是使用可光裂解的ssDNA编码的抗体缀合物来实现具有<75“污染”血细胞的高度纯化的CTC群体。在单细胞条形码芯片上创建增强的聚-L-赖氨酸条形码模式，用于在微室中有效捕获稀有CTC细胞，用于随后的分泌蛋白质分析。用接种到全血中的EpCAM阳性HCT 116细胞对该系统进行了广泛评估和优化。采用患者血液样品来评估该系统用于CTC的分离、纯化和单细胞分泌概况分析的效用。患者血液样品中存在的CTC表现出IL-8和VEGF的高度异质性分泌谱。在平行实验中，发现分泌CTC的数量与基于免疫染色的CTC计数不一致。
Genetic and transcriptional profiling, as well as surface marker identification of single circulating tumor cells (CTCs) have been demonstrated. However, quantitatively profiling of functional proteins at single CTC resolution has not yet been achieved, owing to the limited purity of the isolated CTC populations and a lack of single-cell proteomic approaches to handle and analyze rare CTCs. Here, we develop an integrated microfluidic system specifically designed for streamlining isolation, purification and single-cell secretomic profiling of CTCs from whole blood. Key to this platform is the use of photocleavable ssDNA-encoded antibody conjugates to enable a highly purified CTC population with <75 ‘contaminated’ blood cells. An enhanced poly-L-lysine barcode pattern is created on the single-cell barcode chip for efficient capture rare CTC cells in microchambers for subsequent secreted protein profiling. This system was extensively evaluated and optimized with EpCAM-positive HCT116 cells seeded into whole blood. Patient blood samples were employed to assess the utility of the system for isolation, purification and single-cell secretion profiling of CTCs. The CTCs present in patient blood samples exhibit highly heterogeneous secretion profile of IL-8 and VEGF. The numbers of secreting CTCs are found not in accordance with CTC enumeration based on immunostaining in the parallel experiments.
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可光透明的DNA条形码 - 抗体结合物可以在单个细胞中敏感和多重蛋白质分析。
影响因子：
15
作者：
Agasti, Sarit S.;Liong, Monty;Peterson, Vanessa M.;Lee, Hakho;Weissleder, Ralph
通讯作者：
Weissleder, Ralph
影响因子：
46.9
作者：
通讯作者：
--
影响因子：
46.9
作者：
通讯作者：
--
影响因子：
6.1
作者：
Sheng W;Ogunwobi OO;Chen T;Zhang J;George TJ;Liu C;Fan ZH
通讯作者：
Fan ZH
DOI：
10.1073/pnas.0703478104
发表时间：
2007-06-12
影响因子：
11.1
作者：
Dalerba, Piero;Dylla, Scott J.;Clarke, Michael F.
通讯作者：
Clarke, Michael F.