Recombinant bfgf

Human pre‐implantation embryos were donated from patients who had completed their family after assisted reproduction treatment. Written informed consents were obtained from all the donors recruited and the study protocols were approved by the Institutional Review Boards of the University of Hong Kong/Hospital Authority Hong Kong West Cluster (IRB number: UW 18‐017) and the Council of Reproductive Technology, Hong Kong (research license number: R5004). Cryopreserved day 2 human pre‐implantation embryos were thawed and cultured in G‐1 medium (Vitrolife) for 1 day before transferred to G‐2 medium (Vitrolife) and cultured till morula or early blastocyst stages when the zona pellucida of the embryos was removed using acid tyrode solution (Sigma Aldrich). The human embryos were then placed on the STO feeder cells in hEPSC medium supplemented with 10 ng mL⁻¹ recombinant bFGF (R&D), 20 ng mL⁻¹ recombinant activin A (PeproTech), and 5% FBS (Thermo Fisher Scientific). The embryos showed outgrowth after several days. The resultant cell colonies were dissected into small clusters using glass pipettes and transferred onto new STO feeder cells. After several passages, the colonies were digested with 0.05% Trypsin (Thermo Fisher Scientific) and passaged like other hEPSCs.