Microfluidic Generation of Oxidized Phospholipid Gradients in Supported Lipid Bilayers to Examine Scavenger Receptor Interactions.

Reactive oxygen species (ROS) are responsible for the oxidative truncation of polyunsaturated fatty acids (PUFAs). The products of these reactions have been implicated in many diseases such as cancer and atherosclerosis. As increasing attention is directed toward these oxidized phospholipids (oxPLs), higher throughput methods are needed to examine interactions between oxPLs and scavenger receptors in the immune system. Supported lipid bilayers (SLBs) are a reliable model for lipid membrane-protein interaction studies due to their compositional tunability and ease of integration with a variety of sensing methods. Combining SLBs with microfluidic devices can boost experimental throughput. Herein, we develop a microfluidic gradient generator for creating SLBs that allows for simultaneous analysis of arrayed SLBs while maintaining distinct oxPL concentrations. First, we demonstrate the ability of the system to create linear arrays of SLBs that possess a persistent gradient of oxPL concentrations. The concentration-dependent binding of CD36 scavenger receptor-functionalized vesicles to SLBs containing gradients of the high-affinity CD36 ligand, KDdiA-PC, was observed. Thermal oxidation of the PUFA 1-palmitoyl-2-arachidonoyl--glycero-3-phosphocholine (PAPC) created a mixture of oxidized products to serve as potential CD36 ligands in the assay, with the primary oxidation product being 1-palmitoyl-2-glutaryl--glycero-3-phosphocholine (PGPC). Lastly, simulations showed that the oxidized tails of both KDdiA-PC and PGPC protrude from the membrane and are accessible to scavenger receptor binding. Interestingly, despite its shorter length, the oxidized tail of PGPC extends further from the membrane interface.