Hi,
I would like to remove the various detergents in the SDS sample buffer (Laemmli buffer)
I'm attempting to use a Slide-A-Lyzer cassette from Thermo but the bromophenol dye doesn't seem to be diffusing suggesting the dialysis isn't working as I'd hoped.
Any suggestions on how to improve the dialysis or other methods that may reverse the addition of the SDS buffer?
Thanks in advance
The SDS will not dialyze in any reasonable amount of time from Laemmli sample buffer because its concentration is higher than its critical micellar concentration (CMC). This means that the detergent is in the form of micelles. These micelles are too large to pass through the pores of the dialysis membrane. Diluting the detergent to a concentration below the CMC (about 8 mM) should speed up the dialysis process. As for bromphenol blue, my guess is that the dye is incorporated into the SDS micelles, so it does not dialyze either.
Use urea 8M to disrupt the micelles of detergents and perform multiple times of UF by washing with urea to remove the detergent monomers. Look for FASP applications prior to proteomic analysis to get tips...
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