GST fusion protein quantitation - (Feb/07/2005 )
I am doing some invitro pull down assays using GST-fusion proteins. I want to know how to quatitate these proteins. As these are made in bacteria, I always see some GST in all the fusion protein aliquots when I run them on the gel and stain with coomassie blue. As I want to load equal amount of proteins in the pull down how do I correct for the GST (also other bacterial proteins )present in all of them.
The easiest way to quantify the amount of GST-fusion protein is to compare to BSA standards of known concentrations i.e. 3 microgram, 6 microgram and 10 microgram on a coomassie stained SDS-PAGE gel. Use equal amount of GST-fusion protein even though it’s not completely pure.