Cell recovery assay

Summary

Cell resuscitation is the process of thawing cells frozen in liquid nitrogen and then re-culturing them to restore cell growth.

Principle

The basic principle of cell resuscitation is to melt the cells fast during resuscitation to allow the cells to pass quickly through the most vulnerable -5 - 0°C, preventing the formation of ice crystals inside the cells to cause cell death.

Operation method

cell recovery

Materials and Instruments

Equipment: 37°C water bath, centrifuge, sterile test tubes, and culture flasks.
Reagents:
① RPMI 1640 culture medium containing 20% serum;

Move

The basic process of cell recovery can be divided into the following steps:A Remove the cell ampoule or cryopreservation tube from the liquid nitrogen and place it in a 37°C water bath to thaw rapidly. Remove the cryopreservation tube from the water bath as soon as the ice disappears.B. Scrub the outside of the tube with 70% ethanol to minimize the chance of contamination.C. Aspirate the contents of the cryopreserved tube and slowly add them to a T-25 cell culture flask containing 4-5 ml of culture medium and incubate in a 37℃ cell culture incubator at 5% _CO2, changing the medium on the next day. Alternatively, the cell protectant can be removed by centrifugation and the cells can be cultured again.

Caveat

It is not always possible to achieve 100% success in resuscitation of frozen cells. Failure of resuscitation may be related to the following factors: low cell number or poor cell growth at the time of freezing; contamination of cells with bacteria or mycoplasma; poor storage of liquid nitrogen tanks; change of culture conditions during resuscitation, such as changing fetal bovine serum to calf serum; and inappropriate resuscitation methods.

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