This collection explores the use of pluripotent cells to generate early embryo-like structures in vitro, with the resultant 3D embryo-like structures in vitro phenocopying many of the developmental landmarks and 3D architectures seen normally in vivo. The protocols gathered herein cover diverse aspects of the topic and exemplify some of the key developments and improvements in the field. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.