What the function about Glycine,Tris base and SDS in Western Blot's Running Buffer?
Hi there,
Tris/glycine is the buffering system. In transfer buffer, Tris base is 25mM and glycine 192mM which results in pH around 8.3 whereas the addition of SDS (which is not compulsary) improves protein transfer from PAGE but decreases protein binding to nitrocellulose (methanol or ethanol is more efficient for protein binding to nitrocellulose).
Tris-glycine is buffer for maintaining pH and sds to give overall negative charge to proteins so that proteins move towards anode only
The association of SDS molecules with protein molecules imparts an associated negative charge to the molecular aggregate formed. This negative charge is significantly greater than the original charge of that protein. The electrostatic repulsion that is created by SDS binding forces proteins into a rod-like shape, thereby eliminating differences in shape as a factor for electrophoretic separation in gels
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