Cryopreservation with a twist - towards a sterile, serum-free surface-based vitrification of hESCs.

Human embryonic stem cells (hESCs) play an important role in the fields of regenerative medicine, basic scientific research, tissue engineering and toxicology. Their unique morphology however makes them very sensitive to cryopreservation procedures. We recently introduced a surface dependent, enzyme- and serum-free method for the effective cryopreservation of bulk quantities of hESC colonies using direct immersion into liquid nitrogen (Beier et al., 2011 [5]). However, direct contact with liquid nitrogen risks contamination and cell infection and severely limits clinical application. This work introduces a modified method and a new combined cultivation and cryopreservation device to facilitate the surface dependent vitrification without contact with (possibly unsterile) liquid nitrogen. The technique allows the culture, cryopreservation, storage and post-thawing cultivation in the same device without detaching cell samples from the cultivation surface. Successful vitrification of bulk quantities of hESCs without direct liquid nitrogen contact is an important step towards automated cryopreservation processes for clinical applications of stem cells and other colony forming cell types.