I am currently interested in doing some flow cytometry to accompany confocal protein co-localization studies in order to get a more quantitative measure of the co-localization. The problem is, we need a technique to permeabilize only the plasma membrane and not the nuclear or organelle membranes. Could it be possible for us to use perforin to permeabilize the cells as opposed to methanol or formalin? Any help would be greatly appreciated.
Yes, 0.02 % Triton X-100 apparently selectively permeabilizes cell membrane, leaving the nuclear membrane intact. Here's the reference: Virology. 1988 Jan;162(1):255-9.
Hope this helps.
Agreed. Additionally the more "gentle" per, buffers from a number of companies will do the same thing. Saponin, from memory, will not permeabilise the nucleus.
I agree with both comment. Saponin is a relatively mild detergent that solvates cholesterol present in the plasma membrane. At low concentrations, internal membranes remain intact
Thank you all very much. We will be sure to try both methods and see which one gives us the best result!
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