An alternative to cryopreservation for long-term storage of mammalian cells is being developed by researchers at Oregon State University who reported in PLOS One that their method is a more convenient, less expensive means of preserving mammalian cells for in vitro fertilization, species conservation, cell therapy and more.
Desiccation has many advantages including the potential to enable viable cell preservation at more convenient storage temperatures without the need for liquid nitrogen. Ease of shipping is another benefit.
To achieve stability during storage in the dried state it is necessary to remove enough water that the remaining matrix forms a non-crystalline glassy solid. Thus, the glass transition temperature is a key parameter for design of cell desiccation procedures.
Corresponding author Adam Higgins, associate professor of bioengineering at OSU, and his collaborators studied the glass transition temperatures of various possible desiccation media and how moisture content affected the temperatures. "Glass transition data for complex, multicomponent solutions are relatively scarce," Higgins said. "But as we expected, the glass transition temperatures increased with decreasing moisture content."
"Adding polymers to the solutions increased the glass transition temperature," he said. "Adding penetrating cryoprotectants decreased it." In general, researchers found the polymers they —polyvinylpyrrolidone (PVP) and Ficoll—had a stabilizing effect; adding them to a desiccation medium containing trehalose and phosphate buffered saline increased the glass transition temperature significantly at any moisture content. Adding DMSO, ethylene glycol or propylene glycol to the mixture decreased the glass transition temperature in a manner that depended on concentration.
"For stable storage in the dried state, it's believed the glass transition temperature should be about 50 degrees Kelvin above the storage temperature," Higgins said. "So a desiccation solution containing 0.1 molarity trehalose and 10 percent PVP in phosphate buffered saline would need to be dried to a water mass fraction of about 5 percent for stable storage at room temperature. For storage in a refrigerator, drying to about 9 percent water content would be sufficient. Adding membrane-permeable cryoprotectants can safeguard intracellular organelles but increases the extent of drying necessary."
Results suggest room-temperature storage of a desiccation solution containing 0.25 molarity cryoprotectants is not feasible. But it appears possible for such a solution to achieve stable storage in a standard refrigerator or freezer after drying to water mass fractions of about 3 percent and 6 percent, respectively.