Scaling and automation of a high-throughput single-cell-derived tumor sphere assay chip.
Scaling and automation of a high-throughput single-cell-derived tumor sphere assay chip.
复制标题
DOI：
10.1039/c6lc00778c
复制
发表时间：
2016-10-07
期刊：
影响因子：
6.1
通讯作者：
Yoon E
中科院分区：
文献类型：
--
作者：
Cheng YH;Chen YC;Brien R;Yoon E
近年来的研究表明，肿瘤干细胞样细胞（CSCs）是肿瘤复发和转移的关键亚群。由于表面抗原和酶活性标志物的癌症可塑性，功能性肿瘤球测定是CSC鉴定的有希望的替代方案。为了可靠地量化罕见的CSC（1-5%），需要数千个单细胞悬浮培养物。虽然微流体是处理单细胞的强大工具，但以前的工作提供有限的通量，并且缺乏高通量研究所需的自动数据分析能力。在这项研究中，我们提出了高通量单细胞来源的肿瘤球检测芯片的缩放和自动化，便于在芯片上跟踪高达约10 000个细胞，捕获率约为76.5%。所提出的细胞捕获方案保证了从散装细胞中取样具有代表性的群体。为了分析具有各种荧光强度的数千个单细胞，开发了一种高度适应性的分析程序，用于细胞/球体计数和尺寸测量。使用在聚二甲基硅氧烷（PDMS）上的Pluronic® F108（聚（乙二醇）-嵌段-聚（丙二醇）-嵌段-聚（乙二醇））涂层，创建悬浮培养环境以测试有争议的假设：较大或较小的细胞是否更像干细胞，其由形成单细胞衍生的球体的能力定义。不同的细胞系表现出不同的球体形成率和初始细胞大小之间的相关性，这表明乳腺癌细胞系之间的通路调控的异质性。更有趣的是，通过监测数百个球体，我们确定了球体生长动力学的异质性，表明即使在CSC内也存在细胞异质性。这些初步结果突出了CSC研究中前所未有的高通量和自动化的力量。
Recent research suggests that cancer stem-like cells (CSCs) are the key subpopulation for tumor relapse and metastasis. Due to cancer plasticity in surface antigen and enzymatic activity markers, functional tumorsphere assays are promising alternatives for CSC identification. To reliably quantify rare CSCs (1–5%), thousands of single-cell suspension cultures are required. While microfluidics is a powerful tool in handling single cells, previous works provide limited throughput and lack automatic data analysis capability required for high-throughput studies. In this study, we present the scaling and automation of high-throughput single-cell-derived tumor sphere assay chips, facilitating the tracking of up to ~10 000 cells on a chip with ~76.5% capture rate. The presented cell capture scheme guarantees sampling a representative population from the bulk cells. To analyze thousands of single-cells with a variety of fluorescent intensities, a highly adaptable analysis program was developed for cell/sphere counting and size measurement. Using a Pluronic® F108 (poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol)) coating on polydimethylsiloxane (PDMS), a suspension culture environment was created to test a controversial hypothesis: whether larger or smaller cells are more stem-like defined by the capability to form single-cell-derived spheres. Different cell lines showed different correlations between sphere formation rate and initial cell size, suggesting heterogeneity in pathway regulation among breast cancer cell lines. More interestingly, by monitoring hundreds of spheres, we identified heterogeneity in sphere growth dynamics, indicating the cellular heterogeneity even within CSCs. These preliminary results highlight the power of unprecedented high-throughput and automation in CSC studies.
登录
查看更多内容
影响因子：
6.1
作者：
Frimat, Jean-Philippe;Becker, Marco;West, Jonathan
通讯作者：
West, Jonathan
影响因子：
3.9
作者：
Hellmich, W;Regtmeier, J;Ros, A
通讯作者：
Ros, A
影响因子：
4.6
作者：
Chen YC;Ingram PN;Fouladdel S;McDermott SP;Azizi E;Wicha MS;Yoon E
通讯作者：
Yoon E
DOI：
10.1007/978-3-662-49118-8_1
发表时间：
2016-01-01
期刊：
ESSENTIALS OF SINGLE-CELL ANALYSIS: CONCEPTS, APPLICATIONS AND FUTURE PROSPECTS
影响因子：
--
作者：
Chen, Yu-Chih;Ingram, Patrick;Yoon, Euisik
通讯作者：
Yoon, Euisik
影响因子：
50.3
作者：
Magee JA;Piskounova E;Morrison SJ
通讯作者：
Morrison SJ