Derivation of Human Embryonic Stem Cells

Abstract

Embryonic stem cells (ESCs) represent a mainstay for pluripotent stem cell research and development (R&D) and provide tangible opportunities for clinical translation including cell therapies and drug discovery. Moreover, in spite of the discovery of induced pluripotent stem cells (iPSCs), ESCs are an essential reference point, against which other pluripotent cells are compared. Hence, there is an ongoing need to derive and bank quality-controlled research-grade and clinical-grade ESC lines using established and standardized methods. Here, we provide a concise, step-by-step protocol for the derivation of ESCs from human embryos. While largely based on previously reported method for clinical-grade human ESC (hESC) line derivation, the protocol is suitable for routine application, although adaptable for clinical-compliance.

Pluripotent stem cells: Basic biology or else differentiations aimed at translational research and the role of flow cytometry

“…Eventually, the human ES cell line was derived for the first time by Professor James Thomson's group in the USA [9]. The ESC derivation process essentially involves removing the zona pellucida, then manually picking up the inner cell mass and plating the same onto the feeder cells [10]. The cells are then replenished with ESC media till the cells start forming additional colonies.…”

“…Eventually, the human ES cell line was derived for the first time by Professor James Thomson's group in the USA [9]. The ESC derivation process essentially involves removing the zona pellucida, then manually picking up the inner cell mass and plating the same onto the feeder cells [10]. Interestingly, early ES cell lines growing on feeder cells can be subjected to live-cell staining simultaneously for the pluripotent stem cells (TRA-1-60 Alexa Fluor ® 594) and the feeder cells (CD44 Alexa Fluor ® 488 Conjugate) using such live-cell imaging kits from Thermo Fisher, USA (https://www.thermofisher.com/order/catalog/product/ A25528#/A25528).…”

The roles of TET family proteins in development and stem cells

“…Human ESCs (hESCs), derived from the pre-implantation stage ICM (Crook et al, 2017), differ significantly from mESCs at both molecular and functional levels. They are considered to represent a later state of epiblast development, resembling primed mEpiSCs with regards to their morphology, developmental potency, gene expression and epigenetic modifications (Hackett and Surani, 2014;Rossant, 2015;Smith, 2017).…”