Creating Spatially Addressed Arrays of Planar Supported Fluid Phospholipid Membranes

Abstract

The use of planar supports for presenting large arrays of spatially addressed molecules is one of the most powerful and versatile methods for creating combinatorial libraries.1-3 These systems are starting to form the basis for a new generation of rapid screening assays and sensor devices in the biological and chemical sciences. Extending this approach to supported phospholipid bilayer membranes containing peptides, receptors, and integral membrane proteins is an especially valuable goal because of the ability of these systems to mimic many of the properties of native cell surfaces.4 Addressing biomembrane mimics on planar supports, however, presents unique challenges, as the twodimensional fluidity of the biomembrane must be preserved in many cases for it to function properly.5-7 The bilayer deposition process must take place in an aqueous environment, and the entire system must continue to remain submerged under water to preserve the planar supported structure. Because of this physical constraint as well as the inherent complexities of biomembrane materials, traditional technologies such as light-directed synthesis for addressing peptide or DNA sequences onto solid supports are inherently difficult to apply.1 We have, therefore, employed an alternate approach based upon depositing mesoscopic quantities of aqueous solution onto lithographically patterned hydrophilic surface well plates,8 followed by the immersion of the entire substrate into buffer. This is a general and flexible method for directing chemically distinct phospholipid membranes into individually addressable surface sectors. Previous studies have shown that patterned surfaces allow partitioning of one fluid lipid bilayer from the next.9,10 Molecules within an individual membrane are free to move within the confines of a single partition but do not cross over to a neighboring region. In the experiments presented here, planar borosilicate substrates were partitioned into arrays of micrometer-sized hydrophilic boxes using standard photolithography. Patterning was achieved by exposing the surface to ultraviolet light through a lithographic mask consisting of an array of square boxes. Developing the pattern and cleaning the substrate formed well plates of hydrophilic glass onto which picoliter-sized droplets of liposome solution were placed (Figure 1). The liposomes, which were small unilamellar vesicles (SUVs) of phospholipids, were present at 1 mg/mL concentration in a pH 7.0, 100 mM sodium phosphate buffer solution. Figure 2 shows the epifluorescence image of nine 50 μm × 50 μm well plates that have been addressed with three chemically