Dear Santosh,
Please read the following text:
Cryopreservation
Cryogenic preservation is the current golden standard for long-term cell storage and also allows the generation of (allogenic) master and working cell banks (1). Drawbacks include limited storage capacity due to the use of specialised equipment, use of toxic cryoprotective agents, complicated protocols and relatively high cost.
Slow freezing generally yields over 90 % viability post-thaw (2),(3) but not all cell types respond favourably to cryogenic preservation. For example, poor survival of cryopreserved human embryonic stem cells (hESC) grown as colonies has been reported (4),(5), while a study showed subzero temperatures compromise the functionality of human hepatocytes as cells lose their adherence capabilities and exhibit an altered metabolic profile post-thaw (6).
Additional problems with cryopreservation may include cell damage from intra- and extracellular ice formation and cell dehydration (7), induction of apoptosis upon thawing caused by injury due to osmotic imbalance (8), and the use of animal derived components, such as fetal bovine serum, may introduce contamination in the samples (9).
Finally, dimethyl sulfoxide (DMSO) is the most widely used cryoprotective agent and in the same time represents one of the main problems with cryogenic storage. There are three main reasons for that: At ambient temperatures it is toxic to cells (10) , may induce cell differentiation (11) and patients treated with DMSO-preserved cells can experience adverse reactions (12). Therefore a large portion of the cryogenic research is devoted to finding new safer protective agents.
Nevertheless, cryopreservation, using conventional slow freezing and storage in liquid nitrogen (the vapour phase, -150 °C, or directly submerged in liquid N2, -196 °C) is the most commonly used method to store cells used for research and cryopreservation at -80 or -20 °C is commonly used to store and transfer cells between the manufacturing site and medical facility (13),(14).
The existence of more than one preservation method is due to drawbacks found in such methods. Thus, the possibility of selecting a method from a number of methods eliminates or decreases the risks of cells destruction upon storage.
References:
1-Heidemann, R., Lünse, S., Tran, D. & Zhang, C. Characterization of cell-banking parameters for the cryopreservation of mammalian cell lines in 100-mL cryobags. Biotechnol. Prog. 26, 1154–63 (2010).
2-Kleeberger, C. A. et al. Viability and Recovery of Peripheral Blood Mononuclear Cells Cryopreserved for up to 12 Years in a Multicenter Study. Clin. Diagnostic Lab. Immunol. 6 , 14–19 (1999).
3-Kotobuki, N. et al. Viability and osteogenic potential of cryopreserved human bone marrow-derived mesenchymal cells. Tissue Eng. 11, 663–73 (2005).
4-Li, Y., Tan, J. & Li, L. Comparison of three methods for cryopreservation of human embryonic stem cells. Fertil. Steril. 93, 999–1005 (2014).
5-Xu, X. et al. The roles of apoptotic pathways in the low recovery rate after cryopreservation of dissociated human embryonic stem cells. Biotechnol. Prog. 26, 827–37 (2010).
6-Terry, C., Dhawan, A., Mitry, R. R. & Hughes, R. D. Cryopreservation of isolated human hepatocytes for transplantation: State of the art. Cryobiology 53, 149–59 (2006).
7-Mazur, P. Freezing of living cells: mechanisms and implications. Am. J. Physiol. 247, C125–42 (1984).
8-Mathew, A. J., Van Buskirk, R. G. & Baust, J. G. Improved Hypothermic Preservation of Human Renal Cells Through Suppression of Both Apoptosis and Necrosis. Cell Preserv. Technol. 1, 239–253 (2002).
9-Merten, O. W. Safety issues of animal products used in serum-free media. Dev. Biol. Stand. 99, 167–80 (1999).
10-Hunt, C. J. Cryopreservation of Human Stem Cells for Clinical Application: A Review. Transfus. Med. Hemother. 38, 107–123 (2011).
11-Santos, N. C., Figueira-Coelho, J., Martins-Silva, J. & Saldanha, C. Multidisciplinary utilization of dimethyl sulfoxide: pharmacological, cellular, and molecular aspects. Biochem. Pharmacol. 65, 1035–41 (2003).
12-Cox, M. A., Kastrup, J. & Hrubiško, M. Historical perspectives and the future of adverse reactions associated with haemopoietic stem cells cryopreserved with dimethyl sulfoxide. Cell Tissue Bank. 13, 203–15 (2012).
13-Coopman, K. Large-scale compatible methods for the preservation of human embryonic stem cells: current perspectives. Biotechnol. Prog. 27, 1511–21 (2011).
14-Harel, A. Cryopreservation and Cell Banking for Autologous Mesenchymal Stem Cell-Based Therapies. Cell Tissue Transplant. Ther. 2013, 1–7 (2013).
Hoping this will be helpful,
Rafik
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