The Need for Standardized Cryoprocessing on Preservation Outcomes

Background

Recent literature indicates that the characteristics of ice formation during cryopreservation can impact preservation outcomes in eukaryotic cells. Ice formation can be modulated by various chemical approaches such as anti-freeze proteins or reagents and other physical means such as high temperature ice nucleation and controlled rate freezing. Certain non-adherent human cell lines have poor post-cryopreservation outcomes characterized by significant loss of viability 24 hours post-thaw, which could be due to variation of ice formation characteristics during cryoprocessing. It is a common practice to use passive freezing containers for small scale cryopreservation. However, such freezing conditions have limited scalability and offer very little control over the freezing process. The current work highlights the need for standardization in freezing processes in cryopreserving biomaterials in production environments.

Methods

In this study, preservation outcomes of THP-1 human monocytes isolated from a monocytic leukemia patient (ATCC TIB-202) were compared from multiple controlled rate freezing (CRF) temperature profiles using an identical cryopreservation medium. Cell recovery was measured using standard viability, growth, and metabolic activity. Finally, the thermal characteristics of each CRF profile were analyzed using differential scanning calorimetry.

Results

All recovery metrics recorded were moderately lower when compared to the non-frozen control. Additionally, there was statistically no difference in viability and metabolic activity between the different freezing methods. However, there was a reduction in the variability between the CRF profiles when compared to the passive freezing container. We also report differences in the thermal characteristics between the different freezing profiles.

Conclusions

This work reconfirms that batch-to-batch variability remains a concern in cryopreservation outcome of mammalian cells; standardization of CRF profile could be an important strategy to address the issue. Further investigation is required to identify the exact mechanisms of variability reduction during cryoprocessing.