Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA.
Biotechnol Bioeng. 2010 May 1;106(1):106-18. doi: 10.1002/bit.22661.
We have developed a novel three-dimensional (3D) cellular microarray platform to enable the rapid and efficient tracking of stem cell fate and quantification of specific stem cell markers. This platform consists of a miniaturized 3D cell culture array on a functionalized glass slide for spatially addressable high-throughput screening. A microarray spotter was used to deposit cells onto a modified glass surface to yield an array consisting of cells encapsulated in alginate gel spots with volumes as low as 60 nL. A method based on an immunofluorescence technique scaled down to function on a cellular microarray was also used to quantify specific cell marker protein levels in situ. Our results revealed that this platform is suitable for studying the expansion of mouse embryonic stem (ES) cells as they retain their pluripotent and undifferentiated state. We also examined neural commitment of mouse ES cells on the microarray and observed the generation of neuroectodermal precursor cells characterized by expression of the neural marker Sox-1, whose levels were also measured in situ using a GFP reporter system. In addition, the high-throughput capacity of the platform was tested using a dual-slide system that allowed rapid screening of the effects of tretinoin and fibroblast growth factor-4 (FGF-4) on the pluripotency of mouse ES cells. This high-throughput platform is a powerful new tool for investigating cellular mechanisms involved in stem cell expansion and differentiation and provides the basis for rapid identification of signals and conditions that can be used to direct cellular responses.
我们开发了一种新颖的三维（3D）细胞微阵列平台，可实现干细胞命运的快速高效跟踪和特定干细胞标志物的定量分析。该平台由功能化玻璃载玻片上的小型化 3D 细胞培养阵列组成，可实现空间寻址的高通量筛选。微阵列点样仪用于将细胞沉积到改性玻璃表面上，从而产生由体积低至 60nL 的藻酸盐凝胶斑点包封的细胞组成的阵列。还使用基于免疫荧光技术的方法对细胞微阵列进行了缩放，以原位定量测定特定细胞标志物蛋白水平。我们的结果表明，该平台适合研究小鼠胚胎干细胞（ES 细胞）的扩增，因为它们保持着多能性和未分化状态。我们还在微阵列上检查了小鼠 ES 细胞的神经分化，并观察到神经外胚层前体细胞的产生，其特征是表达神经标志物 Sox-1，还使用 GFP 报告系统原位测量了其水平。此外，通过使用允许快速筛选维甲酸和成纤维细胞生长因子-4（FGF-4）对小鼠 ES 细胞多能性影响的双载玻片系统测试了该平台的高通量能力。该高通量平台是研究干细胞扩增和分化涉及的细胞机制的强大新工具，并为快速鉴定可用于指导细胞反应的信号和条件提供了基础。
