Reviving Cells from Liquid Nitrogen - (Aug/06/2001 )

Hi all....

I need some help in reviving my CHO and HEK cells after storage in liquid nitrogen in 40%serum, 10% DMSO. Any suggestions?

Thanks, Allison

Once you take out the cells from the liquid nitrogen tank.let it thaw in a 37 degrees water bath. Add cells to medium containing 20% serum.Give two washes. Let the cells grow in a medium containing 20% serum for couple of passages.It will work.

The key to reviving cells seems to be:
1) warm them up quickly - I hold them in the water bath and keep shaking. As soon as they thaw then pipette into a tube with medium immediately.
2) don't seed the cells at too low a density.
3) use a higher than normal serum concentration for the seeding (ie 15% rather than 10%)
3) it is generally recommeded that you change the medium as soon as the cells are adherant (the next day) so that the DMSO is removed.

If you are still having problems it may not be the reviving as much as it is the original freezing thats the problem.