在人类发育和疾病相关研究中，更完整的体外模型迫切需要，而器官样体具有巨大潜力。它们复杂的细胞组成使得单细胞测序具有很大的实用性；然而，目前技术限制为少数处理条件，限制了其在器官样体异质性的筛选或研究中的应用。在这里，我们应用sci-Plex，一种基于单细胞组合索引（SCI）的RNA测序（RNA-seq）多重法，对视网膜样体进行分析。我们证明了sci-Plex和10×方法产生了高度一致的细胞类组成，并进一步扩展sci-Plex来分析对关键发育途径进行调节时样体的细胞类组成。通过利用个体样体数据，我们开发了一种测量样体异质性的方法，并确定早期激活Wnt信号在视网膜样体培养中增加了长达6周的视网膜细胞类别。我们的数据显示sci-Plex在分析处理条件对相关人体模型的潜力上可以实现扩大规模。©2023年作者。

With a critical need for more complete in vitro models of human development and disease, organoids hold immense potential. Their complex cellular composition makes single-cell sequencing of great utility; however, the limitation of current technologies to a handful of treatment conditions restricts their use in screens or studies of organoid heterogeneity. Here, we apply sci-Plex, a single-cell combinatorial indexing (sci)-based RNA sequencing (RNA-seq) multiplexing method to retinal organoids. We demonstrate that sci-Plex and 10× methods produce highly concordant cell-class compositions and then expand sci-Plex to analyze the cell-class composition of 410 organoids upon modulation of critical developmental pathways. Leveraging individual organoid data, we develop a method to measure organoid heterogeneity, and we identify that activation of Wnt signaling early in retinal organoid cultures increases retinal cell classes up to 6 weeks later. Our data show sci-Plex's potential to dramatically scale up the analysis of treatment conditions on relevant human models.© 2023 The Authors.