G418 conc for HEK293 cells - (Nov/08/2006 )
What is the right concentration of selection antibiotic(G418) for HEK293 cells?
-minnee-
the best way to know is to determine it yourself.
Treat some non transfected cells with increasing concentrations (around 1 mg/mL)
and determine which concentration gives you the best result
-Missele-
QUOTE (minnee @ Nov 8 2006, 08:14 AM)
What is the right concentration of selection antibiotic(G418) for HEK293 cells?
I've successfully used G418 at a final conentration of 0.8mg/ml on these cells.
-maxi-
800 ug/ml selection, 400ug/ml maintanence
-tap14-
as tap14
-fred_33-
QUOTE (fred_33 @ Nov 9 2006, 01:14 PM)
as tap14
How much time does it take to kill all non transfected cells by adding 800ug/ml?
-minnee-
generally 1-2 weeks.
-fred_33-
QUOTE (fred_33 @ Nov 20 2006, 09:02 PM)
generally 1-2 weeks.
If it takes 1-2 weeks for the non transfected cells to die off, do we change media within that time frame. does dead cells that float around induce any signal to kill off other cells as well? Im just worried that dead cells will give off apoptotic signals to other cells in the same flask.
-dennym85-
That's normal part of the selection process. The true colonies will come out among the dead.
-genehunter-1-
QUOTE (minnee @ Nov 8 2006, 03:14 PM)
What is the right concentration of selection antibiotic(G418) for HEK293 cells?
high conc (>500 µg/ml) are often recommended but you may succeed with ~350 µg/ml; as suggested by other members you have to check the best conc.; use the lowest effective conc, too high conc may force cell cycle arrest for the surviving cells; your transfected gene beside resistance factor may influence cell viability, and therefore selection criteria by G418
-The Bearer-