Cause: After the separation and the fixation lipophilic proteins can get native again even after the gel swam 20 minutes in 20% TCA!
This can only happen if alcoholic staining recipes are used. During this procedure the proteins begin to diffuse, the bands get broader and finally the proteins will leave the gel. Hydrophilic proteins are well stained synchronously.
Remedy: will be the use of non-alcoholic staining recipes.
We suggest to use the Coomassie-Violet staining procedure.
To achieve a clear background as Ampholytes only Servalytes, Sinulytes or SepaLytes should be used. Pharmalytes (GE) give strong background!