Storage temperature of biological sample

  Long-term storage of biological samples typically uses as low a temperature as possible to reduce biochemical reactions within the sample to increase the stability of the various components within the sample. Common storage temperatures for biological macromolecules, cells, tissues and organs are -80 ° C (ultra-low temperature refrigerator), -140 ° C (liquid nitrogen phase or cryogenic refrigerator) and -196 ° C (liquid nitrogen liquid phase), the lower the temperature of the sample The longer the stable storage time. 0 to -60 ° C is the crystallization temperature of water, which easily damages the microstructure of cells and tissues. Generally, this temperature is not used to preserve tissues and cells. Some purified biomacromolecules can be stably stored at 0--60 ° C for a certain period of time, but in the tissue sample, the biological macromolecules are affected by various factors in the cell tissue, and the stability may be significantly reduced, so the sample bank is usually not used. 0 to -60 ° C as the storage temperature.

  -196 ° C sample storage

  -196 ° C is the temperature at which liquid nitrogen volatilizes, so only liquid nitrogen liquid storage technology can reach this temperature. The life activities in the sample are basically stopped at this temperature, and the stability of the sample can be preserved for a long time. It is the most effective method for long-term preservation of cell activity, complex structure and activity of tissues and organs in samples, and is widely recognized. Liquid storage of liquid nitrogen containers requires further protection against cross-contamination between samples compared to other different temperature freezing modes. The National Cancer Institute recommends using a cryotube with a spiral to encapsulate the sample. However, during the cooling process, the sample suddenly drops from the ultra-low temperature freezer (-80 ° C) to liquid nitrogen, causing the shrinkage of the frozen cap and the tube body to be inconsistent, which easily leads to the penetration of liquid nitrogen into the cryotube and thus increases the cross-contamination between the samples. risk. One solution is to heat-shrink each of the cryotubes using a special cryotube sleeve, or use a sealing film to wrap around the interface between the cryotube cap and the body for 2-3 turns. The former method will lengthen the tube and require a higher freezer box. The second method will make the tube body slightly thicker. It is recommended to use a conventional frozen storage box of 10x10 size at the bottom.

  -140 ° C sample storage

  -140 ° C is lower than the glass transition temperature of water (~ -136 ° C), is also the temperature that liquid nitrogen phase and cryogenic refrigerator can reach, the biological activity of the sample in this temperature range is greatly reduced, is to save the sample The ideal temperature for cell activity. Similar to the phase change temperature at which the ice-water mixture can be maintained at 0 °C, the liquid and gaseous nitrogen in the insulated liquid nitrogen container should be maintained at a phase transition temperature of -196 °C. However, in practice, due to the insufficient sealing of the liquid nitrogen container lid, a temperature gradient is formed between the liquid nitrogen level and the liquid nitrogen container can. The National Cancer Institute recommends that the temperature at the mouth of the liquid nitrogen container should be kept below -140 °C. Samples that are not determined for future use should be stored in liquid nitrogen phase mode to protect the cell activity in the tissue.

  The cryogenic refrigerator is electrically cooled, does not require liquid nitrogen, and the stable temperature after filling the sample is usually below -140. Compared with the use of liquid nitrogen, the advantage is that it does not require frequent addition of liquid nitrogen and is easy to maintain. However, the cooling rate of electric cooling is lower than that of liquid nitrogen. Once the container is opened and sampled, it is easy to cause a large range of temperature fluctuations, and the temperature recovery time is relatively long, so it is more suitable for less open and take samples. In addition, the electric refrigeration refrigerator must guarantee power supply. It is recommended to use liquid nitrogen equipment in the event of a power outage to provide backup storage.

  -80 ° C sample storage

  -80 °C is lower than the crystallization temperature range of the more hazardous water, which is also the temperature that can be achieved by the ultra-low temperature refrigerator of common equipment. It is considered based on factors such as ease of operation, storage and cost. The usual temperature for molecular activity. However, it is still inconclusive as to how long the temperature of different biomacromolecules can be maintained. The stability of DNA in tissues can be maintained for several years or longer at -80 °C. However, for RNA, it is easy to be gradually degraded by RNase widely distributed in cells and various tissues. In different cells and tissues, the length of stable storage of RNA is also quite different, but generally it is less than 5 years. In some sensitive experiments, RNA has been degraded in less than one year at -80 °C, so for long-term preservation of RNA activity, it is recommended to use lower temperatures, or use a small sample to extract RNA and remaining samples. Sync storage. Biomacromolecules such as proteins and lipids in other samples can be stored in the sample at -80 ° C, but the duration is different and the stability is gradually attenuated. If it is to protect a specific biological macromolecule known in the sample, a stabilizer for the molecule can also be added. If the biomacromolecules to be preserved in the sample are not determined, it is recommended to use a lower temperature storage. In addition, the current large-scale automated sample access equipment can only be used with -80 ° C ultra-low temperature equipment, and can not be used with liquid nitrogen equipment. UK Biobank in the UK is storing a copy of some samples (~9.5 million) in a working sample at -80 °C, using automated equipment; another copy (~5.5 million) is stored in a liquid nitrogen phase for safety elsewhere. Backup, sample manual access.

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