Polymer-based microfluidic device for measuring membrane protein activities

Abstract

Functional assays of membrane proteins are becoming increasingly important, both in research and drug discovery applications. The majority of current assays use the patch-clamp technology to measure the activity of ion channels which are over-expressed in cells. In future, in vitro assay systems will be available, which use reconstituted membrane proteins in free-standing lipid bilayers suspended in nano-or micrometer-sized pores. Such functional assays require (1) expression, purification and reconstitution of the membrane protein of interest, (2) a reliable method for lipid bilayer formation and membrane protein integration, and (3) a sensitive detection system. For practical applications, especially for automation, the reliable and controllable transport of fluids is essential. In order to achieve a stable free-standing lipid bilayer, a pore diameter in the micro-to nanometer range is essential. Novel microfluidic devices were developed by bonding a thick (300 lm) polyether ether ketone foil, bearing a channel structure, to a thin (12 lm) foil with a micropore of about 10 lm diameter and then utilized for the formation of stable, free-standing lipid bilayers within the pore. A bacterial voltage-gated potassium channel is integrated therein by fusion and the ion channel activity detected by voltage clamp.

Reconstitution of Human Ion Channels into Solvent-free Lipid Bilayers Enhanced by Centrifugal Forces

“…Possible macroscopic events, however, could not be resolved due to the large transient current, which probably resulted from a clamp delay caused by charging across the BLMs with a high capacitance (usually 50-100 pF). Similar slow transients after the voltage steps have also been reported for voltage-gated potassium channels reconstituted in BLM systems (37,42).…”

A label-free microfluidic assay to quantitatively study antibiotic diffusion through lipid membranes

“…Membrane permeation across artificial and cell membranes has been extensively studied in microfluidic platforms 9,10,[27][28][29][30][31] . However, most of the work has concentrated on developing platforms for supported lipid bilayers in microfluidic chambers.…”

Physical modelling, numerical simulation and experimental investigation of microfluidic devices with amorphous thermoplastic polymers using a hot embossing process

“…Injection moulding and hot embossing are the most commonly used replication processes for mass production in a variety of industries. Hot embossing is an ideal method for rapid the prototyping of micro and nano structures into polymers (Hutter et al, 2013), especially for producing delicate microstructures with high aspect ratios (Becker and Heim, 2000). Hot embossing provides several advantages over injection moulding, such as a lower cost for the embossing tool, flexibility in the choice of embossing material and a high replication accuracy for small features.…”