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IMAC of Histidine-Tagged Fusion Proteins 145
Table 3
Wash type Effect Comment
Glycine, arginine,
∼0.5 MNH 4 Cl and
pH 7
Non-amine salts, e.g.,
∼0.5 M–1.0 M NaCl;
in 20 mM Imidazole +
50 mM NaCl pH 7
Non-ionic detergents,
e.g., Triton, Tween no
more than 1% v/v
Chaotropic agents, e.g.,
4 M Urea or 4 M
Guanidine–HCl
Decreasing pH (20 mM)
Mild eluents that
compete for Ni with
histidine
Will disrupt any
non-specific
electrostatic
interactions
Disrupts hydrophobic
interactions
Disrupts the histidine
bond to the IMCC
These are mild eluents
that will not elute the
His-tag protein but may
displace weaker bound
proteins
Such interactions are
common in IMAC
particularly if the
equilibration and wash
steps had