Ultraflat Nickel Substrates for Scanning Probe Microscopy of Polyhistidine-Tagged Proteins

A novel approach to the preparation of ultra-flat Nickel substrates suitable to Scanning Probe Microscopy imaging of immobilized polyhistidine-tagged proteins has been devised. Exploiting freshly cleaved mica, Ni thermal evaporation followed by thermal annealing in vacuum, and the template stripping method, we have obtained Ni substrates with a rms roughness as low as 0.12 nm, which bind readily polyhistidine-tagged proteins, enabling molecular resolution imaging of isolated molecules as well as of molecular sub-monolayers. Protein sample exposure to imidazole causes removal of the adsorbates, confirming the involvement of the poly-histidine tail in protein surface immobilization.