I am attempting GST-pull down assay for finding unknown interacting partners using GST-fusion protein. Normally bait control, prey control, resin control are taken but I am planning to take also a vector control, i.e., only GST as GST itself can bind to several interactors to give a false positive result.
In my view, a GST-only control is the best negative control. Just replace your GST-tagged protein of interest with free GST and keep all other pulldown conditions the same. This will allow you to control for all sources of unspecific binding, namely the resin and GST itself. Cheers,
GST pulldown is a valuable technique to investigate if two proteins interact. If you would like to determine the binding affinity in addition, I would recommend to use MicroScale Thermophoresis.
Here is a three minutes video to explain the technique:
http://www.nanotemper-technologies.com/news-events/news/article/watch-a-3-minutes-movie-to-explore-microscale-thermophoresis-mst/
For further questions, please feel free to contact me via [email protected]!
- Badges
- Science topic
- Similar topics
- Chemistry
- General Biochemistry
- Proteomics