Serum starving cells - why? - (Apr/03/2007 )

Hello,

I am new to cell culture and I been coming across protocols that call for serum-starving cells before transfection or cell activation. What is the reason/purpose for serum starving cells?

Hi
there is many reasons to starve cells before experiments, for example you put all the cells into G0 phase and with serum they are in all phases of cell cycle.
If you want to transfect cells with some lipid reagent like lipofectamine you should use serum free medium to decrease toxicity, I think there are some components of serum that normaly don´t cross the membrane, but can enter cells with transfection reagent and are toxic in this high concentrations.
There are also other reasons, like serum itself contains some enzymes, like MMPs and if you want to measure this activity you must eliminate the contribution of the serum.
Maybe others can think of some more reasons.
H.

I have done transfections with various reagents and serum starving is not necessary. One can get high transfection rates without this step.

The primary reason is empirical--it just works.

A good example is of stimulation with growth factors. A wide variety of cells type actually will not show much response (as measured by activation of downstream components of GF receptors, for example) if they are stimulated with factor after being grown in normal medium (I've personally found this to be the case for insulin stimulation, often). This is presumably due to negative-feedback mechanisms that serve to downregulate the constant signals cells are receiving from stimulation in their medium (e.g. serum proteins/growth factors)--many times serum starving seems to ablate the negative feedback and allows you to actually see robust responses. So, basically, it's one of the things people always try when they can't see signaling responses in cells.

The other reason, as mentioned before, is to halt proliferation of cells. This is done for various reasons--some people will culture MEFs (primary fibroblasts) at certain times in low serum to stop them from dividing and using up their "finite" allotted doublings before senescence. The other reason is to prevent cells from overgrowing the plate; if you grow cells to, say 70% confluence but you're not ready to do a procedure on them for some reason, their proliferation can be slowed down by removing the serum.

Serum contains loads and loads of natural ligands for all kinds of receptors, obviously to get that signaling down to basal, you need to use serum free conditions...

D