Early post-implantation human embryonic development has been challenging to study due to both technical limitations and ethical restrictions. Proper modeling of the process is important for infertility and toxicology research. Here we provide details of the design and implementation of a microfluidic device that can be used to model human embryo development. The microfluidic human embryo model is established from human pluripotent stem cells (hPSCs), and the resulting structures exhibit molecular and cellular features resembling the progressive development of the early post-implantation human embryo. The compartmentalized configuration of the microfluidic device allows the formation of spherical hPSC clusters in prescribed locations in the device, enabling the two opposite regions of each hPSC cluster to be exposed to two different exogenous chemical environments. Under such asymmetrical chemical conditions, several early post-implantation human embryo developmental landmarks, including lumenogenesis of the epiblast and the resultant pro-amniotic cavity, formation of a bipolar embryonic sac, and specification of primordial germ cells and gastrulating cells (or mesendoderm cells), can be robustly recapitulated using the microfluidic device. The microfluidic human embryo model is compatible with high-throughput studies, live imaging, immunofluorescence staining, fluorescent in situ hybridization, and single-cell sequencing. This protocol takes ~5 d to complete, including microfluidic device fabrication (2 d), cell seeding (1 d), and progressive development of the microfluidic model until gastrulation-like events occur (1–2 d).

由于技术限制和伦理限制，早期植入后人类胚胎发育的研究一直具有挑战性。该过程的正确建模对于不孕症和毒理学研究很重要。在这里，我们提供了可用于模拟人类胚胎发育的微流体装置的设计和实施细节。微流控人类胚胎模型是从人类多能干细胞 (hPSCs) 建立的，所得结构表现出类似于早期植入后人类胚胎逐渐发育的分子和细胞特征。微流体装置的分隔配置允许在装置的指定位置形成球形 hPSC 簇，使每个 hPSC 簇的两个相对区域能够暴露于两种不同的外源化学环境。在这种不对称的化学条件下，几个早期的植入后人类胚胎发育标志，包括外胚层的腔发生和由此产生的羊膜腔前，双极胚囊的形成，以及原始生殖细胞和原肠胚细胞（或中内胚层细胞）的规格，可以使用微流体装置进行稳健的概括。微流控人类胚胎模型与高通量研究、实时成像、免疫荧光染色、荧光原位杂交和单细胞测序兼容。该协议需要约 5 天才能完成，包括微流控设备制造 (2 d)、细胞播种 (1 d)、