High Content Image Analysis of Spatiotemporal Proliferation and Differentiation Patterns in 3D Embryoid Body Differentiation Model

The 3D embryoid body (EB) differentiation model is a promising tool for fundamental cell biology and drug discovery studies assessing the compound effects on mammalian and human development. This 3D cell model allows for analyzing spatiotemporal changes during morphogenesis and differentiation. A combination of confocal microscopy with high content image analysis (HCIA) can significantly improve the study of spatiotemporal patterns of early embryonic lineages and compound efficacy and toxicity testing by enhancing the identification and quantification of various cell types. HCIA can be used to assess the EB architecture through quantitative and qualitative characteristics, such as viability and apoptosis, identification, localization, ratio and timing for various types of early embryonic cells, dimensions of compartments of proliferating and differentiating cells, changes in the size and shape of EBs, and translocation of individual cells and cell layers. This chapter describes a comprehensive framework for HCIA for 3D EB differentiation model that allows investigators to analyze EB growth, differentiation, and morphogenetic dynamics.