Enhancement of cell cryopreservation with glycolipids

Inventors

Campbell, Lia H. • Brockbank, Kelvin G M

Assignees

Lifenet Health

Abstract

Living cellular material may be preserved by incubating the cellular material in a culture medium containing at least one glycolipid, and then subjecting the cellular material to a preservation protocol.

Core Innovation

The disclosure relates to preserving living cellular material by exposing the cellular material to a cryoprotectant composition containing at least one anti-freeze glycolipid and a cryoprotectant for 6-72 hours. The cellular material is selected from keratinocytes, hepatocytes, and cardiac myocytes, and the medium is formulated as a cryoprotectant composition having the cryoprotectant present in an amount from 0.05 M to 11 M.

The antifreeze glycolipid includes glycolipids isolated from freeze-tolerant insects and plants, including Tipula trivittat, Ceruchus piceus, Solanum dulcamara, Dendroides canadensis, and Cucujus clavipes. The disclosure further describes AFGLs having a xylomannan/β-mannopyranosyl-(1↔β-xylopyranose backbone bearing a lipophilic moiety, and states that such antifreeze glycolipids increase post-thaw cell viability and improve survival and proliferation.

After the exposure period, the method subjects the cellular material to a preservation protocol that cools from 37°C to a sub-zero temperature of the medium at a cooling rate of 10 to 30°C per minute, followed by cryopreservation at a cryopreservation temperature of about ≤-80°C or less. The disclosure emphasizes that cell viability after completion of the preservation protocol is at least 60%.

Claims Coverage

The claims cover a method for preserving selected living cellular material using a cryoprotectant composition that combines an antifreeze glycolipid with a cryoprotectant, followed by a defined cooling and cryopreservation protocol that yields at least 60% post-preservation cell viability. The independent claim specifies four inventive features.

Overall, claim coverage centers on combining at least 0.1 ppm anti-freeze glycolipid with a cryoprotectant for 6-72 hours in keratinocytes, hepatocytes, and cardiac myocytes; using cryoprotectant present at 0.05 M to 11 M; applying cooling at 10-30°C per minute from 37°C to a sub-zero temperature followed by cryopreservation at about ≤-80°C or less; and achieving at least 60% cell viability after completion.

Stated Advantages

Documented Applications