Cryopreservation of animal cells

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BACKGROUND

To maintain the cell culture stock, we store integrated viable cells in cryovials in liquid N₂ tank for the application in future. It saves all expensive chemical resources for the cell culture also.

In this experiment, we aimed to cryopreserve healthy and integrated cells in liquid N₂ for future usage.

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REQUIREMENTS

 

Sample:      Animal cells in a culture plate (confluent stage).

Apparatus: 15ml centrifuge tube,

Cryovials,

Serological Pipette,

Centrifuge with rotor.

Chemicals:  1X HBSS,

Cell culture media,

0.5% trypsin solution,

Serum,

DMSO.

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PROCEDURE

Media for the cryopreservation

Take out the confluent cell plate from the incubator discard the culture media and wash cells 1X HBSS and add 3 ml 0.5% trypsin solution (per 60mm petri dish), incubate at 37°C for 3-5 min and wait until all the cells are detached from the plate. Add 5-7 ml of cell culture media and mix it carefully further try to dissociate all the cell clumps in the solution. Centrifuge it at 300x g for 3-5 min and discard the supernatant further add the cryopreservation media and mix properly and keep in to the -20°C immediately for 1-2 hrs then you can transfer in to liquid N₂ tank for long time storage.¹

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CONCLUSION

This experiment was concluded with the cryopreservation of live and integrated cells for the future application.

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REFERENCES

1. Kumar A, Mishra HK, Dwivedi P, Subramaniam. Secreted trophic factors of Human umbilical cord stromal cells induce differentiation and neurite extension through PI3K and independent of cAMP pathway. Annals of neuroscience. 2015;22(2):97-106.