Characterization of human embryonic stem cells in animal component-free medium

Clinical use of human embryonic stem cells (ESCs) as a raw material requires good manufacturing practice-compliant axillary materials such as culture medium. To this end, animal components should not be used and contamination of virus/bacteria/fungus and allergens are a concern. In addition, animal components such as albumin and fetal bovine serum pose difficulties such as a lot-to-lot variation. However, only a limited number of animal component-free media have been developed to date. In this study, we investigated whether SEES2 ESCs can be stably propagated for 16 passages (54 population doublings) over a period of 60 days in a newly established Stem-Partner® ACF medium. SEES2 ESC maintained their intact karyotype, i.e. 46,XX, and their undifferentiated phenotypes after long-term culture. An in vitro differentiation assay revealed that SEES2 ESCs exhibited multipotency, i.e. endodermal, mesodermal and ectodermal differentiation. Subcutaneous implantation of SEES2 ESCs generated mature teratomas without malignant transformation. These results show that SEES2 ESCs in the Stem-Partner® ACF medium can be used to establish master cell banks for future regenerative medicine as well as other ESCs in the previously reported culture medium. ### Competing Interest Statement Hiroshi Miyajima and Tetsuji Sasaki are employees of Kyokuto Pharmaceutical Industrial Co., Ltd., and Yoshiko Abe is an employee of SEKISUI MEDICAL CO., LTD.