Format

Send to

Choose Destination
Biotechnol J. 2018 Feb;13(2). doi: 10.1002/biot.201700384. Epub 2017 Oct 30.

A Simple and Robust Method for Culturing Human-Induced Pluripotent Stem Cells in an Undifferentiated State Using Botulinum Hemagglutinin.

Author information

1
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan.
2
Laboratory for Infection Cell Biology, International Research Centre for Infectious Diseases, Research Institute for Microbial Diseases, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan.
3
Department of Bacteriology, Graduate School of Medical Sciences, Kanazawa University, 13-1 Takara, Kanazawa, Ishikawa, 920-8640, Japan.

Abstract

Clinical and industrial applications of human-induced pluripotent stem cells (hiPSCs) is hindered by the lack of robust culture strategies capable of sustaining a culture in an undifferentiated state. Here, a simple and robust hiPSC-culture-propagation strategy incorporating botulinum hemagglutinin (HA)-mediated selective removal of cells deviating from an undifferentiated state is developed. After HA treatment, cell-cell adhesion is disrupted, and deviated cells detached from the central region of the colony to subsequently form tight monolayer colonies following prolonged incubation. The authors find that the temporal and dose-dependent activity of HA regulated deviated-cell removal and recoverability after disruption of cell-cell adhesion in hiPSC colonies. The effects of HA are confirmed under all culture conditions examined, regardless of hiPSC line and feeder-dependent or -free culture conditions. After routine application of our HA-treatment paradigm for serial passages, hiPSCs maintains expression of pluripotent markers and readily forms embryoid bodies expressing markers for all three germ-cell layers. This method enables highly efficient culturing of hiPSCs and use of entire undifferentiated portions without having to pick deviated cells manually. This simple and readily reproducible culture strategy is a potentially useful tool for improving the robust and scalable maintenance of undifferentiated hiPSC cultures.

KEYWORDS:

E-cadherin disruption; botulinum hemagglutinin; human-induced pluripotent stem cells; removal of deviated cells; undifferentiated state

PMID:
29027750
DOI:
10.1002/biot.201700384
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Wiley
Loading ...
Support Center