Manstein Felix, Halloin Caroline, Zweigerdt Robert
Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiac, Thoracic-, Transplantation and Vascular Surgery, REBIRTH-Cluster of Excellence, Hannover Medical School, Hannover, Germany.
Methods Mol Biol. 2025;2924:59-71. doi: 10.1007/978-1-0716-4530-7_5.
This paper describes a detailed protocol for human pluripotent stem cells (hPSCs) cultivation as matrix-free cell-only aggregates in defined and xeno-free culture medium in stirred tank bioreactors (STBRs). Starting with a frozen stock pre-expanded on conventional culture dishes (2D), the ultimate process is performed in 150 mL culture scale in stirred tank bioreactors (3D) and is designed to produce up to 500 million pluripotent hPSC within 7 days. The culture strategy includes perfusion-based cell feeding facilitating process control, automation, and higher cell yields. Ultimately, this detailed protocol describes an important step for generating a defined starting cell population for directed lineage differentiation and subsequently fueling human cell-based assays and regenerative medicine approaches.
本文描述了一种在搅拌罐生物反应器（STBRs）中，于限定的无动物成分培养基中，将人多能干细胞（hPSCs）作为无基质纯细胞聚集体进行培养的详细方案。从在传统培养皿（二维）上预扩增的冻存细胞开始，最终过程在150毫升培养规模的搅拌罐生物反应器（三维）中进行，旨在7天内生产多达5亿个多能hPSC。培养策略包括基于灌注的细胞补料，有助于过程控制、自动化和更高的细胞产量。最终，此详细方案描述了为定向谱系分化生成限定起始细胞群体的重要步骤，进而推动基于人细胞的检测和再生医学方法。
