Long-term microcarrier suspension cultures of human embryonic stem cells

The conventional method of culturing human embryonic stem cells (hESC) is on two-dimensional (2D) surfaces, which is not amenable for scale up to therapeutic quantities in bioreactors. We have developed a facile and robust method for maintaining undifferentiated hESC in three-dimensional (3D) suspension cultures on matrigel-coated microcarriers achieving 2- to 4-fold higher cell densities than those in 2D colony cultures. Stable, continuous propagation of two hESC lines on microcarriers has been demonstrated in conditioned media for 6 months. Microcarrier cultures (MC) were also demonstrated in two serum-free defined media (StemPro and mTeSR1). MC achieved even higher cell concentrations in suspension spinner flasks, thus opening the prospect of propagation in controlled bioreactors. (C) 2009 Elsevier B.V. All rights reserved.