Functionalisation of human chloride intracellular ion channels in microfluidic droplet-interface-bilayers

Abstract

Profiling ion flux through human intracellular chloride ion channels using live-cell based techniques, such as patch-clamp electrophysiology, is laborious and time-consuming. The integration of scalable microfluidic systems with automatable protocols based on droplet-interface-bilayers (DIBs) within which ion channels are incorporated circumvents several limitations associated with live-cell measurements and facilitates testing in controllable in vitro conditions. Here, we have designed and tested novel microfluidic layouts for the formation of arrays of DIBs in parallel and developed the first example of a miniaturised, DIB-based, fluorescence assays for Cl− fluxing, allowing the investigation of the functional properties of the human chloride intracellular ion channel 1 (CLIC1). The microfluidic protocols relied on passive geometries for droplet pairing and DIB formation. Using recombinantly expressed CLIC1, we identified the best conditions to maximise protein integration into a lipid bilayer and the oligomerisation of the protein into functional ion channels. Finally, CLIC1 ion channel functionality was assessed relative to α-Haemolysin into microfluidic DIBs using the same Cl− fluxing assay.