u/Clear-Negotiation796

Iam trying to study if my compound could inhibit nfkb activation when stimulated with LPS, that is at the end measuring p-NF-kB p65 levels, Now with my western blot I can see a clear total NFkB Levels, but p-NF-kB is little ambiguous. Now here my protocol details:

cells: Thp1 dervied macrophages

lps concentration: 20ng/ml

stimulation time: 30min

fbs concentration: regular 10% in RPMI media,

Are these factors correct, many say i have to reduce fbs concentration or even reduce it to zero, or should i increase lps concetration like 100ng/ml and what about stimulation time

Any suggestions would be great