History and Process

Cell cryopreservation is the process of protecting and preserving viable cells by cooling the cells to very low temperatures, typically −130°C to −196°C using liquid nitrogen. The first experiments on freezing cells started in 1949 by Polge, Smith and Parkes who succeeded in cryopreserving ram semen. Since then, significant advances have been made in cell cryopreservation techniques and applications. Today, a wide variety of cells including hematopoietic stem cells, skin cells, embryos and tumor cells are routinely cryopreserved for future therapeutic and research applications.

The basic steps involved in cell cryopreservation include slow cooling and rapid thawing. In slow cooling, the cells are slowly cooled at a controlled rate of around 1°C/minute in the presence of cryoprotectant agents which prevents ice crystal formation inside cells during freezing. Then, the frozen cells are placed in liquid nitrogen for long term storage and thawed rapidly when required by removing from liquid nitrogen and placing in warm media. This prevents formation of ice crystals inside cells during thawing. Successful cryopreservation requires optimizing all parameters like type and concentration of cryoprotectants, cooling and thawing rates for different cell types.

Applications in Cell Banks and Biotechnology

Cell cryopreservation offers major advantages for establishing cell banks of novel cell lines developed for bioproduction, tissue engineering and drug screening purposes. It helps companies maintain repository of well-characterized master and working cell banks for consistent commercial manufacturing over several years. Cryopreservation also enables distribution of cells globally to various research and production facilities. This has aided rapid expansion of cell-based biotechnology industry worldwide.

Cell banks play a key role in stem cell therapy sector. Many private and public cord blood banks worldwide collectively store millions of cryopreserved umbilical cord blood stem cell units that can be used as an alternative to bone marrow transplantation for the treatment of over 80 diseases. Cord blood stem cells have additional advantages like immediate availability and reduced risks of transplant rejection. This promotes promising clinical applications in treating blood disorders.

Cryopreservation helps store cells for future experiments and applications in regenerative medicine. For example, adipose-derived stem cells extracted from donated fat tissues are cryopreserved for potential use in cartilage and bone repair, wound healing, cardiac repair and preventing urinary incontinence. Similarly, skin cell banks help address chronic skin wounds and burns by providing stored keratinocytes and fibroblasts for autologous skin grafting procedures after thawing.

Applications in Pharmaceutical Industry

Cell cryopreservation plays a crucial role in drug discovery, toxicity testing and production of biotherapeutics in the pharmaceutical industry. Cryopreserved cells provide standardization and consistency required for reliable screening of drug candidates as well as toxicity and efficacy evaluation of new drugs during preclinical studies.

In addition, cryopreserved cell lines ensure uninterrupted supply of cells to product development teams engaged in process development, analytical method validations and clinical trial material production related to different biologics like monoclonal antibodies, vaccines, cytokines and recombinant proteins. This allows companies to bank multiple vials of well characterized cells at different passage numbers for emergency back-up supply and future projects.

Cryopreservation offers major advantages over traditional 2D cell culturing methods. It helps to eliminate continuous subculturing, which ensures genetic and phenotypic stability and homogeneity of cultured cells. It also reduces time, effort and cost related to repeated cell procurement, authentication and characterization. Over the decades, consistent progress in optimizing key parameters like choice of cryoprotectants has enhanced viability and growth recovery in various types of adherent and suspension cell lines post-thaw. This promotes wide applications in biomanufacturing.

Challenges and Advancements

While cells from many different sources have been successfully cryopreserved, certain cell types like neurons, hepatocytes, pancreatic islet cells presenting unique sensitivity to freezing damage, ice crystal formation, osmotic and oxidative stresses during cryopreservation. This necessitates further research to achieve higher post-thaw viabilities and functionalities.

Advancements in field of vitrification offer promising alternatives to conventional slow freezing methods for better preservation of such metabolically active and fragile cell types. Vitrification involves ultrarapid cooling rates using high concentration of cryoprotectants to avoid ice crystal formation entirely and form an amorphous glass-like solid matrix. This newer approach has enabled enhanced viability and functionality after cryopreservation and banking of cells like oocytes and ovaries. Continuous progress in understanding molecular mechanisms impacting cell survival during freezing and thawing will help address technical challenges and support wider applications.

Cell cryopreservation is an indispensable technology supporting cell repositories, biological research and commercial manufacturing by ensuring consistent supply of viable cells for extended periods. Continuous innovations aimed at improving cell recovery, functionality and banking of broader range of cell types will further catalyze applications in diverse sectors of regenerative medicine, biotechnology and biopharmaceutical industry.

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