When is the best time to freeze cells for long-term storage?

Freezing cells for long-term storage is most effective when performed during the log phase at approximately 80% confluency. At this stage, cells are actively dividing and are in their most healthy and metabolically active state, which enhances their viability upon thawing. Cells at this density have a balanced ratio of cell density and nutrient availability, making them more resilient during the freezing process.

During freezing, cells are subjected to stress due to the formation of ice crystals, which can damage cellular structures. Freezing them at the optimal confluence level ensures that a sufficient number are alive and will recover well after being thawed. This is critical because successful long-term storage depends on the ability of the cells to survive and maintain their functional characteristics post-thaw.

In contrast, freezing at stationary phase or during differentiation may result in lower cell viability and functionality due to the physiological changes that occur during these states. Cells in stationary phase are not dividing and may have already started to undergo senescence, and differentiated cells are often more specialized, making them less capable of re-entering the cell cycle after thawing. Thus, the choice of freezing cells at 80% confluence during log phase supports optimal recovery and functionality in long-term storage.