Kaori Sugihara

Liposome and Lipid Bilayer Arrays Towards Biosensing Applications

Sensitive and selective biosensors for high-throughput screening are having an increasing impact in modern medical care. The establishment of robust protein biosensing platforms however remains challenging, especially when membrane proteins are involved. Although this type of proteins is of enormous relevance since they are considered in >60% of the pharmaceutical drug targets, their fragile nature (i.e., the requirement to preserve their natural lipid environment to avoid denaturation and loss of function) puts strong additional prerequisites onto a successful biochip. In this review, the leading approaches to create lipid membrane-based arrays towards the creation of membrane protein biosensing platforms are described. Liposomes assembled in micro- and nanoarrays and the successful set-ups containing functional membrane proteins, as well as the use of liposomes in networks, are discussed in the first part. Then, the complementary approaches to create cell-mimicking supported membrane patches on a substrate in an array format will be addressed. Finally, the progress in assembling free-standing (functional) lipid bilayers over nanopore arrays for ion channel sensing will be reported. This review illustrates the rapid pace by which advances are being made towards the creation of a heterogeneous biochip for the high-throughput screening of membrane proteins for diagnostics, drug screening, or drug discovery purposes.

Artificial Bacterial Flagella for Remote‐Controlled Targeted Single‐Cell Drug Delivery

We hypothesized that ABFs can be functionalized with liposomes, thus allowing these micromachines to per-form biological or biomedical tasks in a remotely con-trolled fashion. To test this hypothesis fl uorescently labeled liposomes and calcein-loaded liposomes were adsorbed on the surface of ABFs. The adsorption of liposomes on the ABFs was confi rmed by quartz crystal microbalance with dissipation monitoring (QCM-D) and fl uorescence recovery after photobleaching (FRAP). The liposome-functionalized ABFs were then placed in contact with cells in vitro and the uptake of calcein (a model water soluble drug) by cells was monitored using fl uorescence microscopy. The fabrication of ABFs was followed as described ear-lier by Tottori et al.

Electrochemical plasmonic sensors

The enormous progress of nanotechnology during the last decade has made it possible to fabricate a great variety of nanostructures. On the nanoscale, metals exhibit special electrical and optical properties, which can be utilized for novel applications. In particular, plasmonic sensors including both the established technique of surface plasmon resonance and more recent nanoplasmonic sensors, have recently attracted much attention. However, some of the simplest and most successful sensors, such as the glucose biosensor, are based on electrical readout. In this review we describe the implementation of electrochemistry with plasmonic nanostructures for combined electrical and optical signal transduction. We highlight results from different types of metallic nanostructures such as nanoparticles, nanowires, nanoholes or simply films of nanoscale thickness. We briefly give an overview of their optical properties and discuss implementation of electrochemical methods. In particular, we review studies on how electrochemical potentials influence the plasmon resonances in different nanostructures, as this type of fundamental understanding is necessary for successful combination of the methods. Although several combined platforms exist, many are not yet in use as sensors partly because of the complicated effects from electrochemical potentials on plasmon resonances. Yet, there are clearly promising aspects of these sensor combinations and we conclude this review by discussing the advantages of synchronized electrical and optical readout, illustrating the versatility of these technologies.

A gigaseal obtained with a self-assembled long-lifetime lipid bilayer on a single polyelectrolyte multilayer-filled nanopore.

A lipid bilayer with gigaohm resistance was fabricated over a single 800 nm pore in a Si3N4 chip using 50 nm liposomes. The nanopore was prefilled with a polyelectrolyte multilayer (PEM) that triggered the spontaneous fusion of the lipid vesicles. Pore-forming peptide melittin was incorporated in the bilayer, and single channel activities were monitored for a period of 2.5 weeks. The long lifetime of the system enabled the observation of the time-dependent stabilization effect of the melittin open state upon bias application.

Electrical polarization of nuclear spins in a breakdown regime of quantum Hall effect

The authors have developed a method for electrical polarization of nuclear spins in quantum Hall systems. In a breakdown regime of odd-integer quantum Hall effect (QHE), excitation of electrons to the upper Landau subband with opposite spin polarity dynamically polarizes nuclear spins through the hyperfine interaction. The polarized nuclear spins in turn accelerate the QHE breakdown, leading to hysteretic voltage-current characteristics of the quantum Hall conductor.

Directed self-assembly of lipid nanotubes from inverted hexagonal structures

Conventional lipid-tube formation is based on either a tube phase of certain lipids or the shape transformation of lamellar structures by applying a point load. In the present study, lipid blocks in inverted hexagonal phase made of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) were shown to protrude lipid nanotubes upon a fluid-dynamic flow on polyelectrolyte-functionalized surfaces in physiological buffer solution. The outer diameter of the tubes is 19.1 ± 4.5 nm and their lengths are up to several hundred micrometers. The method described enables the alignment and patterning of lipid nanotubes into various (including curvy) shapes with a microfluidic system.

A universal method for planar lipid bilayer formation by freeze and thaw

A procedure based on freezing and thawing was developed to induce the rupture of adsorbed lipid vesicles on solid surfaces into supported lipid bilayers (SLBs). The SLB assembly exploits the phase transition of both lipids and water during freezing. It enables SLB formation independent of the type of substrates and lipids as long as the vesicles spontaneously adsorb onto the surface. The created SLB is a single bilayer, and has a diffusion coefficient of (0.6–4) × 10−8 cm2 s−1 on TiO2, which is in the same range as the SLBs formed by conventional techniques. The presented approach has the advantages of both the Langmuir–Blodgett method (the versatility in the selection of lipids and substrates) and vesicle fusion (self-assembly) at the same time.

The Resistance of Polyelectrolyte Multilayers in a Free-Hanging Configuration

The resistivity ρ(PEM) of polyelectrolyte multilayers (PEMs), PEI(PSS/PAH)(24), PEI(PGA/PAH)(12), PEI(HA/PLL)(12) and PEI(PSS/PLL)(12), in a free-hanging configuration was estimated combining electrochemical impedance spectroscopy (EIS) and atomic force microscopic (AFM) images. Surprisingly, the obtained value of several kΩcm is at least 6 orders of magnitude lower than that reported previously, where the resistivity was determined in the conventional PEM-on-electrode system. The significant discrepancy indicates the unexpectedly low electrical PEM resistance in the absence of redox-active ions and the sensitivity limitation in the conventional system.

Combined Electrical and Optical Characterization of Polydiacetylene

Polydiacetylene (PDA) is a conductive polymer that has mechanochromism. When the polymer is exposed to mechanical stresses, change in temperature (thermochromism), pH (ionochromism), and so forth, the structural perturbation can be seen by the change in its color. Although it presents interesting electrical and optical properties, the relationship between these signals has rarely been investigated. We studied the correlation between the electrical conductivity and the absorption spectra of PDA. Upon UV irradiation, PDA absorption spectra presented a blue shift, which coincided with the decrease in the electrical conductivity.

Artificial tubular connections between cells based on synthetic lipid nanotubes

Tunneling nanotubes (TNTs) have become a major topic of interest as a form of intercellular communication due to their recent discovery. However, research on this subject has often suffered from a lack of controllability in the generation of the nanotubular connections. In this work, we demonstrate a simplified approach to selectively create a direct nanotubular connection between eukaryotic cells by manually manipulating self-assembling lipid nanotubes (LNTs) from inverted hexagonal-phase lipid blocks. The technique requires minimal instrumentation for creating the LNT connection between cells compared to conventional approaches. Based on the diffusion of fluorescent lipids from LNTs into cell membranes (D = 0.032 ± 0.003 μm2 s−1), the probability of observing membrane fusion between LNTs and cell membranes was estimated as 30%. Among these cell–LNT junctions the resulting structure is open-ended roughly 75% of the time, as evidenced from observations of the diffusion of a water-soluble dye between two cells connected with this nanotubular structure.