proliferation assay - just a couple of questions (Oct/07/2004 )
Let me give you the gist of what's goin' on.
I'm doing a proliferation assay on MCF-7 cells (I haven't killed them yet, YAY ). It's done in triplicate, with a control, an inhibitor, and a drug, looking at the cells over a week. I was wondering, does the media the cells are grown in, need to be changed throughout the assay, 'cause by day 7, the well looks iffy, though the cells seem quite healthy and happy...the little grots. and how is this done?
Another thing, i get the point of a proliferation assay...but after a northern blot has to be done...WHY? This is something shocking, but i don't get how proving that the drug causes an increase in cells can be supported in a northern. i'm really stupid. and tired, , and can't make the link. Please help me. I'll be your best friend.
Possibly increase in proliferation is supposedly correlated with increase in RNA, because more protein is needed to be synthesized new cells?
What's your Northern for? What's your probe? Maybe it's some gene that is involved in proliferation? In that case it would be just another way to double-check your results. I guess.
As for your media, if your cells are happy you're probably ok. But if you feel uneasy wash out your wells as normal, add fresh mediaum, drug and whatever else is in your asssay- this of course would mean you are using up more reagents. But then aagain, how stable are your chemicals in tissue culture conditions- if not very stable you'll have a better chance that your cells have been subjevted to the effect of your chemicals for the entire week.