Screening ion-channel ligand interactions with passive pumping in a microfluidic bilayer lipid membrane chip

Abstract

We describe a scalable artificial bilayer lipid membrane platform for rapid electrophysiological screening of ion channels and transporters. A passive pumping method is used to flow microliter volumes of ligand solution across a suspended bilayer within a microfluidic chip. Bilayers are stable at flow rates up to $0.5 ll/ min. Phospholipid bilayers are formed across a photolithographically defined aperture made in a dry film resist within the microfluidic chip. Bilayers are stable for many days and the low shunt capacitance of the thin film support gives lownoise high-quality single ion channel recording. Dose-dependent transient blocking of a-hemolysin with b-cyclodextrin (b-CD) and polyethylene glycol is demonstrated and dose-dependent blocking studies of the KcsA potassium channel with tetraethylammonium show the potential for determining IC 50 values. The assays are fast (30 min for a complete IC 50 curve) and simple and require very small amounts of compounds (100 lg in 15 ll). The technology can be scaled so that multiple bilayers can be addressed, providing a screening platform for ion channels, transporters, and nanopores. V C 2015 AIP Publishing LLC.

Characterization of the Prokaryotic Sodium Channel NavSp Pore with a Microfluidic Bilayer Platform

“…Fig 1(C) is a photograph of the bilayer aperture with the integrated Ag/AgCl electrode and the flow channel. Bilayers were made by painting lipid across the aperture and are stable for flow rates up to 0.4 μl/min [ 11 ]. Compared with traditional bilayer electrophysiology systems, both the bilayer capacitance and shunt capacitance is reduced so that higher bandwidth, low noise electrical recordings are possible [ 12 ].…”

“…The glass chips contain integrated Ag/AgCl electrodes together with edge connectors that make direct contact to the ASICs, significantly reducing contact resistance and noise. The bilayers are formed across miniature apertures made in an epoxy photoresist and the design provides fluid access on both sides, via a microfluidic channel ( Fig 1C and 1D ) that provides a simple means of quickly introducing compounds to the bilayer using passive pumping from droplets, eliminating syringe pumps and associated tubing [ 11 ]. In this work, the ion channels are inserted into the bilayer by the addition of proteoliposomes which spontaneously fuse with the lipid bilayer, leading to incorporation of the pores.…”

Artificial Cell Membrane Systems for Biosensing Applications

“…As a result of the intensive development of membrane platforms, recent studies have presented signal features not only of prokaryotic ion channels but also of eukaryotic channels, including those of human origin. ,, Typically, time-courses of ionic current are observed at different applied voltages. , However, for practical and common use of these systems, the signal data will have to be accumulated and carefully validated by comparison with previous patch-clamp data, in terms of channel conductance, open probability, and dwell time distributions. For ligand-gated ion channels, dose-dependent inhibition/activation characteristics were also investigated to verify whether the system would be feasible as a screening procedure and whether the incorporated ion channels were intact within the artificial membrane platforms. , For screening tests, device configuration was studied in order to integrate a fluidic channel for ligand exchange. , A parallel screening system would be an alternative solution and would significantly enhance data throughput (Figure d) . Devices for mechanosensitive channels are ongoing .…”

“…37,38 For screening tests, device configuration was studied in order to integrate a fluidic channel for ligand exchange. 39,40 A parallel screening system would be an alternative solution and would significantly enhance data throughput (Figure 1d). 18 Devices for mechanosensitive channels are ongoing.…”