Permeabilisation for PI staining in FACS? - (Mar/12/2007 )

Hi Everybody
I want to measure transfection efficiency using FACS. I transfect cells with plasmid containing sequence fof GFP. Then I want to fix them and stain with PI.
I am going to fix cells with 3,7% PFA.
Can anybody tell me if I need permeabilisation for PI staining.
Could you please tell me also more detailed protocol: time of fixation for example and at which stage and in which solution I can put the samples to the fridge.
Thanks a lot in advance

Hi,

Why do you want to use PI? If you want to stain for dead cells, you cannot use fixed cells.
Only living cells are able to 'pump out' PI.

Just do not fix them, take some cell to the FACS, add PI and see what you have.

I know, there are some cases where you have to fix the cells (hopefully not in your case), but the you can not use PI (to stain dead cells).

Actually I wanted to stain all the cells with PI and see what part of the total population expresses GFP