Hi, I have procured N2a cells, from a lab that had maintained the cells in DMEM-HG w/ Glutamax (supplement of Glutamine).

I am basically trying to capture increasing death pattern with increasing conc of stressor. I have actually tried growing and experimenting in both kinds of media:

1. ATCC/NCCS recommended media: (MEM-w/ NEEA and Pyruvate, 5.5mM Glucose) + 10% FBS + Pen/Strep (w/o Glutmax but 2mM basal Glutamine)

2. Parental media: (DMEM: HG (25mM) w/ Glutamax)

In both cases, when experiment is done w/in 16-19 PN (PN15 is the lowest I have) it gives a moderately reproducible data. Actually I should not be calling reproducible, because with every increase in PN, the cell death rate is decreasing for any particular conc of stressor.

What's the way out?

Any help will be much appreciated!