Here we discuss strategies for the design and quantitative control of protein-functionalized solid supported lipid bilayers (SLBs) using the reversible interaction between polyhistidine tags and nickel-chelating lipids. SLBs are synthetic, biomimetic surfaces which maintain the two-dimensional fluidity of biological membranes. They are uniquely well-suited for studying an ever-growing array of cell-surface phenomena and membrane-mediated signaling events, but their experimental flexibility is limited by the type and number of proteins with which they can be functionalized.

Nickel-chelating lipids show promise as a general strategy for the attachment of histidine-tagged proteins to SLBs. However, the nickel-histidine interaction is reversible and previously groups have reported substantial desorption rates that would severely limit the system's utility in biological experiments.

Using fluorescence microscopy we show that despite reported claims, nickel-chelating lipids are suitable anchors for attaching systems of proteins to SLBs, and that protein surface density is tunable, allowing for the assembly of surfaces with a variety of protein densities using a single concentration of nickel-chelating lipid. Adsorbed species exist in two distinct populations, one with a surface residence time on the order of minutes, and the other on the order of hours. We can control the density of each species by adjusting the loading conditions, allowing ad hoc design of protein-functionalized SLB surfaces.

The observed behavior is well-described by a two-state binding model in which protein is either monovalently-bound through a single histidine residue, or polyvalently-bound, the latter having a significantly longer residence time on the surface. We discuss the implications of this two-state model with respect to efficient use of protein in SLB-based experimental design, and highlight applications investigating the mechanisms of T-cell recognition in the immunological synapse.