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Open Access Publications from the University of California

Block Copolymer Membranes for Protein Nanopore-Based Biosensing

  • Author(s): Rofeh, Justin
  • Advisor(s): Theogarajan, Luke S
  • et al.
Abstract

Model membranes are synthetic structures that mimic cell membranes. They may be used for basic research or as part of an engineered device. Phospholipids are typically used to make model membranes, but they may be replaced by amphiphilic block copolymers that are more physically and chemically stable, more tunable, and less costly. In this work, we explore the use of block copolymer membranes for biosensing applications. Specifically, we target the goal of nanopore DNA sequencing using the protein nanopore MspA in block copolymer membranes.

As a first step, we extended previous work on MspA in a block copolymer membrane by translating it to an array of apertures using a microfluidic painting technique. We found that the use of a volatile solvent in a microfluidic channel caused variations in membrane properties with corresponding undesirable variations in MspA behavior. Additionally, we used this method to demonstrate DNA translocation through MspA in block copolymer membranes. We selected droplet interface bilayers (DIBs) to investigate which membrane properties are responsible for variations in MspA behavior. However, we found that our aqueous block copolymer DIBs behave differently than any previously studied DIB, necessitating the use of new techniques to characterize their behavior. Key differences include reduced bilayer stability due to poor packing of the monolayer and slow equilibration kinetics. We addressed the poor bilayer stability by establishing a technique for manually packing monolayers prior to bilayer formation. We addressed the slow equilibration kinetics by establishing a new method for measuring monolayer and bilayer tensions in DIBs using droplet shape analysis. While droplet shape analysis is typically used for single droplets with axial symmetry, our method works even for DIBs with asymmetric volumes and tensions. Another advantage is that the method can determine monolayer and membrane tensions in real time, independent of the applied voltage. The method may be used for polymer and phospholipid DIBs alike. The work presented in this thesis contributes to the understanding of amphiphilic block copolymer behavior and will aid in the optimization of membrane properties for biosensing applications.

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