1.
The cell is packaged by dry ice. When receiving the cell, please make sure that the vial is still frozen. If there is cell thawing in the tube, please take photo before experiment or storage.
2.
If immediate culturing is not intended, the cryovial(s) must be stored in liquid nitrogen (-196°C) or at least at -80°C after arrival.
If immediate culturing is intended, please follow these instructions:
3.
Quickly thaw by rapid agitation in a 37℃ water bath within 45-90 seconds. The water bath should have clean water containing an antimicrobial agent. As soon as the sample has thawed, remove the cryovial from the water bath.
Note: A small ice clump should still remain and the vial should still be cold.
From now on, all operations should be carried out under aseptic conditions.
4.
Transfer the cryovial to a sterile flow cabinet and wipe with 70% alcohol. Carefully open the vial and transfer the cell suspension into a 15 ml centrifuge tube containing 9 ml of cell complete medium (room temperature or 37℃).
5.
In order to reduce cell damage, add 1ml of cell complete medium into cryovial, slightly pipette, then use a pipettor to add 1 ml of suspension into the centrifuge tube. Resuspend the cells carefully. Centrifuge at 300×g for 3 min and discard the supernatant. The centrifugation step may be omitted, but in this case the remains of the freeze medium have to be removed 24 hours later.
6.
Resuspend the cells carefully in 10ml fresh cell culture medium and transfer them into one or two T25 cell culture flasks.
