Ruggedized Chip-Based Lipid Bilayer Arrays with Permeable multi-MAC Support

Abstract

Planar lipid bilayers play a central role in nanopore sensing and ion-channel electrophysiology. However, bilayer fragility often limits their application. Previously, we enhanced the mechanical properties of planar lipid bilayers with a single-layered, molecularly thin, minimal actin cortex (MAC). To further increase bilayer robustness, we are developing multiple layered MAC structures (multi-MACs) that can be chemically cross-linked to arbitrary thicknesses, without blocking diffusive access to the outer leaflet of the bilayer. In this work, we describe the multi-MAC formation process in the context of a bilayer array on a fluidic chip, as well as on glass supported bilayers. We report on permeability tests at both the single-molecule and macroscopic levels that show multi-MACs do not significantly block access to the bilayer. We discuss how permeability is achieved by controlling the openness of the F-actin network, which can be tailored using the density of linkage sites, the electrostatic interactions between linkers and filaments, and specific F-actin growth and deposition techniques. We also present results from single-molecule electrical and optical measurements that characterize the multi-MAC structure, permeability, and the approximate pore size of the multi-layered network.