According to the literature relevant to my project, I am to incubate prepared membrane samples with the chemical cross-linkers Diamide, EGS (both following 30min DDT treatment), and DMP for defined times - this I have noted as 30mins.
Would there be a suggested time of incubation with samples containing whole cells (E.coli) as opposed to the membrane samples? I can only assume it must be longer/ of higher concentration?
Do you know if this crosslinker can pass through the e. coli cell wall?
this has got trial and error written all over it I'm afraid. Have you thought about making spheroplasts first to remove the cell wall - plenty of protocols will do that for you.
You can play around with:
different concentrations and/or different times of reaction...
As the "proximity" of the crosslinker depends also of your protein of interest
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