Universal red blood cells (RBCs) differentiated from O-negative human induced pluripotent stem cells (hiPSCs) could find applications in transfusion medicine. Given that each transfusion unit of blood requires 2 trillion RBCs, efficient bioprocesses need to be developed for large-scale in vitro generation of RBCs.

We have developed a scalable suspension agitation culture platform for differentiating hiPSC-microcarrier aggregates into functional RBCs and have demonstrated scalability of the process starting with 6 well plates and finally demonstrating in 500 mL spinner flasks. Differentiation of the best-performing hiPSCs generated 0.85 billion erythroblasts in 50 mL cultures with cell densities approaching 1.7 × 10⁷ cells/mL. Functional (oxygen binding, hemoglobin characterization, membrane integrity, and fluctuations) and transcriptomics evaluations showed minimal differences between hiPSC-derived and adult-derived RBCs.

The scalable agitation suspension culture differentiation process we describe here could find applications in future large-scale production of RBCs in controlled bioreactors.

从 O 阴性人类诱导多能干细胞 (hiPSC) 分化而来的通用红细胞 (RBC) 可用于输血医学。鉴于每个输血单位需要 2 万亿个红细胞，因此需要开发有效的生物过程来大规模体外生成红细胞。

我们开发了一个可扩展的悬浮搅拌培养平台，用于将 hiPSC 微载体聚集体区分为功能性红细胞，并证明了该过程的可扩展性，从 6 孔板开始，最终在 500 mL 旋转烧瓶中进行演示。表现最佳的 hiPSC 的分化在 50 mL 培养物中产生了 8.5 亿个成红细胞，细胞密度接近 1.7 × 10 ^{7 个}细胞/mL。功能（氧结合、血红蛋白表征、膜完整性和波动）和转录组学评估显示 hiPSC 衍生的和成人衍生的 RBC 之间的差异很小。

我们在此描述的可扩展搅拌悬浮培养分化过程可以在未来在受控生物反应器中大规模生产红细胞中找到应用。