Cryopreservation of Human Embryonic Stem Cells with a New Bulk Vitrification Method1

Abstract

To increase the manipulation efficiency and storage capability of vitrified human embryonic stem cells, a new bulk vitrification method was established using transformed cryovials. This method vitrified a large number of cell clumps, as opposed to those cryopreserved by a slow-freezing method with conventional cryovials at one time (round). After warming, vitrified human embryonic stem cells exhibited a much higher survival rate than the slow-freezing cells. The vitrified stem cells continued to express markers of pluripotency and formed teratomas in mice with severe combined immunodeficiency, confirming the pluripotency of vitrified-warmed human embryonic stem cell clumps. The new bulk vitrification method is superior to and more practical than the open pulled straw vitrification method and the slow-freezing method for the cryopreservation of human embryonic stem cells.

Cryopreservation of Human Pluripotent Stem Cells: Are We Going in the Right Direction?

“…In addition, comparable results to those obtained for the classical vitrification method were reported for the recovery rate, the degree of differentiation and the maintenance of the pluripotency of the surviving cells (Li et al, 2008a). A refinement of this technique, using a cryovial fitted with stainless steel mesh, produced similar results (Li et al, 2010a). Although this method is easy and efficient to perform it still presents the limitation of direct contact of the cells with liquid nitrogen increasing the possibility of contamination and cell infection.…”

Vitreous Cryopreservation of Human Preantral Follicles Encapsulated in Alginate Beads with Mini Mesh Cups

“…This homemade cryo-container was then sterilized and ready for use [29]. The encapsulated preantral follicles to be vitrified were placed into the cup-shaped stainless steel mesh (Fig.…”

Cryopreservation of Human Stem Cells for Clinical Application: A Review

“…When compared to hES cells recovered by the OPS method, no significant differences were detected in either the rate of re-attachment or in the degree of differentiation (measured at day 7) nor in the pluripotency of the surviving cells. A refinement of this technique, using customised cryovials fitted with stainless steel mesh, produced similar results [135]. Whilst both methods allow for an increase in the quantity of hES cells preserved at any one time, neither method is suitable for automation and commercial scale-up nor do they overcome the potential contamination problems associated with direct exposure to liquid nitrogen.…”