Protein Extraction from SDS-PAGE - (Jul/22/2008 )
I have a mixture of a few proteins and I need to apply purified fractions in cell culture. So I planned to do SDS-PAGE, to stain the bands with Coomassie-blue and to cut them out subsequently. But with protein extraction out of polyagrylamid gels I do not have any experience and I did not find any protocol. So I do not have any idea how to proceed. Do you by chance have any suggestions?
Very hard to get protein out of SDS-PAGE. And your proteins will be denatured with SDS, likely not active. Choices are
- electroelution : ok for small proteins
- Prep-Cell I think was the name, from BioRad, where you actually run the proteins out of the gel, collect fractions.
I would suggest to separate them by liquid chromatography, try size exclusion, and then you can concentrate the fractions before adding to cells.