low viability of stably transfected cell line from cryostock - (Aug/23/2006 )
I stably transfected mouse embryonic fibroblasts using G418 as selection. I selected clones after single cell cloning in the presence of selection media with G418. I made freeze downs of my new clones. But when I went to thaw the new clones, I see very little viability. I made the freeze downs in the similar way I would for the parent cell line the only difference being that I used the selection media. I dont know what could be wrong? I am wondering should i be using just regular media W/o G418, in that case would i be losing my construct?
freeze thaw cycle is not that good for any living cells/bacterias....
hence the fact that viability after a thaw is poor is a general fact. The difference between cell lines may come from the fact certain cells divide at higher ratio and show more cels on the plate when you're looking your culture for check.