Once considered “old school,” cryopreservation is going through another scientific and technological development growth phase. Today’s new cryopreservation processes and products promise to deliver disruptive technologies to discovery science, stem cell research, diagnostics and personalized medicine; forever changing the future of cell preservation strategies.
Cell preservation has evolved from simple, often poorly annotated private collections of clinical specimens to highly organized and well-annotated collections maintained by commercial and not-for-profit organizations. Currently, the focus of cell preservation strategies is from government and international agencies on the activities of biobanks, specifically on the need to adopt best practices and to provide scientific, legal and ethical guidelines for the biopreservation industry. Cell preservationists still focus on fulfilling the immediate post-thaw viability needs of today’s researchers.
Use of cryopreserved cells is on the rise and this increased utilization drives biobanking demand and production, both in sample quality and utility. Tomorrow’s researchers will need more than high immediate post-thaw cell viability – they will require samples that deliver high cell recovery and recovered products that are physiologically and biochemically identical to their pre-freeze states at the genomic, proteomic, structural, functional and reproductive levels. Biobanks must adapt their strategies and protocols to address the immediate and future needs of their clients.
Hypothermic storage uses a metabolic suppression strategy to maintain biological material, using cold to cause reversible depression of cellular functions. Existing research documents support the protective effects of cold but science has only recently begun to shed light on the biological consequences of cold exposure.
Modern hypothermic and cryopreservation techniques limit ischemia, hypoxia and other negative effects associated with cell and tissue harvest and isolation in a way that increase storage intervals. Ultra-low temperatures stop a cell’s metabolism to allow an indefinite storage period. This approach is currently effective only for single cell suspensions and a few simple tissues, as “cryoninjury” limits results of cryopreservation of whole organs or complex tissues. Hypothermic storage remains the most effective preservation strategy for complex biologics.
Whole organ preservation requires cold perfusion upon harvest, a method that is far superior to warm perfusion and storage but one that offers limited preservation times of only hours to days. Research advanced the ability to recognize cellular responses to stress and expanded knowledge of tolerable limits. Preservation solutions will continue to evolve, even as the development of complex tissue/organ preservation approaches face significant limitations. Advancements in the understanding of the molecular response of cells to cold support targeted approaches that extend preservation intervals and produce a high quality product.
Cryopreservation of biologics currently relies on low temperatures to provide “on demand” access. While controlling osmotic flux, the formation of ice and associated stresses are the standard of practice today, research suggests molecular-based stress response pathways and the control of these pathways is critical to optimal cryopreservation outcome. The molecular aspects of cryopreservation influences cell death but also have a long-term effect on biochemical pathways and cellular functionality after thawing.
A paradigm shift in cryopreservation strategy is necessary to provide high quality samples capable of fulfilling future research needs. The shift must include structural methodologies to prevent ice-related damage and overcome significant cell death after thawing.
Standardization of the isolation steps and the addition of anti-stress agents are key elements of cold chain optimization strategy. The process of cryopreservation can cause molecular-based cell death after thaw. From the moment a technician lifts cells from culture or a surgeon excises tissue, oxidative stress starts compromising normal cell physiology. Pre-freeze manipulation can trigger cell death cascades post-thaw.
Several strategies, used individually or in combination, can suppress cryopreservation-induced molecular-based cell death. Using of cryopreservation media that features intracellular-like ion distribution, impermeants to protect the sample from osmotic extremes and appropriate organic buffers is one such strategy, as is using various free radical scavengers to address sample oxidation. The use of targeted apoptotic inhibitors is also an effective strategy to reduce post-thaw cell death. Another strategy may be to adjust the timing of the addition of complex cryopreservation media according to cell type.
Advances of Cell Preservation on a Molecular Scale
Cryopreservation affects the molecular response of cells. The control and direction of this response has a significant effect on the outcome. The new breed of cell preservation strategies creates a molecular-based foundation and provides a new direction for innovative research, technology development and scientific procedures. Expanding the knowledge and use of cryopreservation helps improve overall efficacy and outcome.
The aim of all biobanks is to preserve biological specimens that researchers will later utilize to support the development of knowledge related to disease control or other relevant purpose. There is a large variety of available biobanking sample types and methodologies, and each is well suited and qualified for its individual purpose. Unfortunately, products that are “fit for purpose” today may not serve other unanticipated purposes in the future. To remain useful in the decades ahead, modern biobanking preservation strategies must accurately predict future needs. Tomorrow’s successes depend on today’s methodologies of preservation that, unfortunately, tend to yield samples of limited utility.
The paradigm shift must focus on organization and funding, and on the locations and procedures involved with biospecimen collections. As the demand increases for a steady supply of high quality and clinically annotated biospecimens, new public-private partnerships and collection sites are necessary to support future sustainability and increase biospecimen availability.
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