Measuring receptor expression by FACS - interference with trypsin exposure? (May/17/2006 )
I wanted to study the expression of a cytokine receptor express at low level. However, I've some problem to detect its expression by FACS (2 different Ab tested at optimal concentrations) (I previously confirmed its expression at the mRNA level).
My question: what are the optimal conditions using adherent cells? Does trypsin exposure impaires receptor expression? What about the use of versene?
Thanks for responses..
Don't know about the use of versene...
But: trypsin will cleave your receptor... So you need your cells time to re-express it...
A trick some people at a different lab in our department do is to trypisn the cells, then incubate them for 2 hours in PBS with 2-5% FCS and then stain them with the antibody's. In PBS the cells won't attach but will (due to the presence of serum maybe) express the receptors.
try to detach the cells with some EDTA (2-5mM)