Microfluidic fabrication of addressable tethered lipid bilayer arrays and optimization using SPR with silane-derivatized nanoglassy substrates

We report the microfluidic fabrication of robust and fluid tethered bilayer arrays within a poly(dimethylsiloxane) (PDMS) chip, and demonstrate its addressability and biosensing by incorporating the GM1 receptor into the bilayer framework for detection of cholera toxin. Rapid optimization of the experimental conditions is achieved by using nanoglassified surfaces in combination with surface plasmon resonance. The ultrathin glassy film on gold mimics glass surfaces employed in microfluidics, allowing real-time monitoring of multiple assembly steps and therefore permitting rapid prototyping of microfluidic arrays. The tethered bilayer array utilizes a covalently immobilized biotinylated protein for generation of well-defined capture zones where a streptavidin link is employed for the immobilization of biotinylated vesicles. Fusion of captured vesicles is accomplished using a concentrated PEG solution, and the lateral diffusion of the tethered bilayer membrane is characterized by fluorescence recovery after photobleaching methods. The tethered membrane arrays demonstrate marked stability and high mobility, which provide an ideal host environment for membrane-associated proteins and open new avenues for high-throughput analysis of these proteins.