Question

What component of the elution buffer causes column-bound PFU to release from the Ni-NTA? Why does this component have this effect on the PFU-bound resin?

Answer 1

Imidazole is a component of the elution buffer that causes the release of column-bound PFU from the Ni-NTA. Imidazole competes with His-tag for the binding to Ni-NTA. As a result, at high concentration (i.e., 150-300 mM), imidazole is a competitive inhibitor of the interaction between histidine and Ni-NTA resin leading to the elution of column-bound PFU from Ni-NTA resin.