Alginate Hydrogel Microencapsulation Inhibits Devitrification and Enables Large-Volume Low-CPA Cell Vitrification

Cryopreservation of stem cells is important to meet their ever-increasing demand by the burgeoning cell-based medicine. The conventional slow freezing for stem cell cryopreservation suffers from inevitable cell injury associated with ice formation and the vitrification (i.e., no visible ice formation) approach is emerging as a new strategy for cell cryopreservation. A major challenge to cell vitrification is intracellular ice formation (IIF, a lethal event to cells) induced by devitrifi cation (i.e., formation of visible ice in previously vitrified solution) during warming the vitrified cells at cryogenic temperature back to super-zero temperatures. Consequently, high and toxic concentrations of penetrating cryoprotectants (i.e., high CPAs, up to approximate to 8 M) and/or limited sample volumes (up to approximate to 2.5 mu L) have been used to minimize IIF during vitrifi cation. It is revealed that alginate hydrogel microencapsulation can effectively inhibit devitrifi cation during warming. The data show that if ice formation were minimized during cooling, IIF is negligible in alginate hydrogel microencapsulated cells during the entire cooling and warming procedure of vitrifi cation. This enables vitrifi cation of pluripotent and multipotent stem cells with up to approximate to 4 times lower concentration of penetrating CPAs (up to 2 M, low CPA) in up to approximate to 100 times larger sample volume (up to approximate to 250 mu L, large volume).