Spelke Dawn P, Ortmann Daniel, Khademhosseini Ali, Ferreira Lino, Karp Jeffrey M
Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
Methods Mol Biol. 2011;690:151-62. doi: 10.1007/978-1-60761-962-8_10.
Embyroid body (EB) formation is a key step in many embryonic stem cell (ESC) differentiation protocols. The EB mimics the structure of the developing embryo, thereby providing a means of obtaining any cell lineage. Traditionally, the two methods of EB formation are suspension and hanging drop. The suspension method allows ESCs to self-aggregate into EBs in a nonadherent dish. The hanging drop method suspends ESCs on the lid of a dish and EBs form through aggregation at the bottom of the drops. Recently, alternative methods of EB formation have been developed that allow for highly accurate control of EB size and shape, resulting in reproducibly produced homogeneous EBs. This control is potentially useful for directed differentiation, as recent studies have shown that EB size may be a useful determinant of the resulting differentiated cell types. One particular approach to generate homogeneous EBs utilizes nonadhesive microwell structures. The methodology associated with this technique, along with the traditional approaches of suspension and hanging drop, is the focus of this chapter.
拟胚体（EB）形成是许多胚胎干细胞（ESC）分化方案中的关键步骤。拟胚体模仿发育中胚胎的结构，从而提供了一种获得任何细胞谱系的方法。传统上，形成拟胚体的两种方法是悬浮法和悬滴法。悬浮法使胚胎干细胞在非贴壁培养皿中自我聚集成拟胚体。悬滴法将胚胎干细胞悬浮在培养皿盖上，拟胚体通过液滴底部的聚集而形成。最近，已开发出形成拟胚体的替代方法，这些方法能够高度精确地控制拟胚体的大小和形状，从而可重复地产生均匀的拟胚体。这种控制对于定向分化可能是有用的，因为最近的研究表明，拟胚体大小可能是所产生的分化细胞类型的一个有用决定因素。一种产生均匀拟胚体的特定方法利用了非粘性微孔结构。与该技术相关的方法，以及悬浮法和悬滴法等传统方法，是本章的重点。
