Optimal protocols for restripping membranes

[agador]

Hey I've got a lot of membranes to reprobe and when I've done it in the past its been quite inconsistent in terms of the quality of the second exposures. Is there anyone out there that may have some pointers about protocols to keep the signal clear and the background low. We have some very limited (therefore) precious samples and need to do many strips.

Anthony

[danfive]

I used Restore Plus (Pierce) ---> http://www.piercenet.com/products/br...e-e7f50837e9e9

Usually it takes 15 minutes (but some Ab bind very strong and will require two 15 minute incubations).
Another tip is to optimize Antibody concentrations, the less Ab the more effective the stripping process.

[agador]

danfive, what is the difference between the product you recommend and a stripping buffer of beta-mercaptoethanol, SDS, Tris HCL pH=6.8. Do you find from your experience that Restore Plus works better?

[danfive]

Originally Posted by agador

danfive, what is the difference between the product you recommend and a stripping buffer of beta-mercaptoethanol, SDS, Tris HCL pH=6.8. Do you find from your experience that Restore Plus works better?

The difference will be that this product won't smell like beta-merc, and probably won't need as much washes like SDS. Those are obvious matters of convenience and not performance. You can definitely make your own.

[agador]

danfive,
Can you reuse Restore Plus after stripping a membrane or do you have to discard it after each use... it might be expensive after a while

[danfive]

Discard it.