Cell-Derived Vesicles with Increased Stability and On-Demand Functionality by Equipping Their Membrane with a Cross-Linkable Copolymer

Cell-derived vesicles retain the cytoplasm and much of the native cell membrane composition. Therefore, they are attractive for investigations of membrane biophysics, drug delivery systems, and complex molecular factories. However, their fragility and aggregation limit their applications. Here, the mechanical properties and stability of giant plasma membrane vesicles (GPMVs) are enhanced by decorating them with a specifically designed diblock copolymer, cholesteryl-poly[2-aminoethyl methacrylate- b -poly(ethylene glycol) methyl ether acrylate]. When cross-linked, this polymer brush enhances the stability of the GPMVs. Furthermore, the pH-responsiveness of the copolymer layer allows for a controlled cargo loading/release, which may enable various bioapplications. Importantly, the cross-linked-copolymer GPMVs are not cytotoxic and preserve in vitro membrane integrity and functionality. This effective strategy to equip the cell-derived vesicles with stimuli-responsive cross-linkable copolymers is expected to open a new route to the stabilization of natural membrane systems and overcome barriers to biomedical applications

Wrapping of Microparticles by Floppy Lipid Vesicles

Lipid membranes, the barrier defining living cells and many of their sub-compartments, bind to a wide variety of nano- and micro-meter sized objects. In the presence of strong adhesive forces, membranes can strongly deform and wrap the particles, an essential step in crossing the membrane for a variety of health and disease-related processes. A large body of theoretical and numerical work has focused on identifying the physical properties that underly wrapping. Using a model system of micron-sized colloidal particles and giant unilamellar lipid vesicles with tunable adhesive forces, we measure a wrapping phase diagram and make quantitative comparisons to theoretical models. Our data is consistent with a model of membrane-particle interactions accounting for the adhesive energy per unit area, membrane bending rigidity, particle size, and vesicle radius

Wrapping of Microparticles by Lipid Membranes

Lipid membranes play an essential role in both the morphology and functionality of living cells. They not only delineate the inside and outside of all eukaryotic cells but also define a number of organelles inside these cells. Cells come into contact with many different objects like particles, pathogens or smaller compartments confined by a lipid membrane, such as vesicles. It is thus essential to understand how membranes interact with such objects and how this interaction is impacted by the physical properties of the membranes. In this thesis, a novel model system to study particle-membrane interactions in proposed. This model system consists of micron-sized particles and giant unilamellar vesicles (GUVs) that interact via nonspecific depletion interactions. In the first part of this thesis, we address the specific conditions for the wrapping of single particles by the membrane. We found three distinct regimes of wrapping: While at low particle-membrane adhesion energies, the membrane does not wrap around the particle, at high adhesion energies particles are spontaneously wrapped. At intermediate adhesion energies particles are wrapped only when an external force is applied. This regime is stabilized by membrane curvature. In the second part of this thesis the dynamics of particle wrapping is investigated in the spontaneous wrapping regime at high adhesion energies. The wrapping dynamics are found to be largely independent of the particle size. Dissipation at the contact line appears to be the dominant process determining the wrapping velocity. However, we observe discrepancies in the experimentally observed wrapping velocities when compared to expected wrapping velocities in a quasistatic model of the system. These discrepancies suggest another process at play on top of dissipation at the contact line. A possible explanation of this phenomena is that the membrane is deformed on a size and timescale in these experiments that the membrane shear viscosity comes into play. The findings of this thesis provide new experimental insights into particle-membrane interactions. Particularly, the proposed novel model system allows for a large number of new experiments offering deeper understanding of membrane mechanics, how cells interact with their environment and how cells shape themselves

Understanding phase transformations in steels using modern electron microscopy techniques

The advantages and limitations of electron back-scattering diffraction coupled with energy dispersive X-ray spectroscopy and of transmission Kikuchi diffraction in relation to the in-depth characterisation of steel microstructures containing phases with similar lattice parameters and/or precipitates are discussed. An in-house developed EBSD map post-processing methodology provided insights into the mechanisms of bainite formation

Dynamics of spontaneous wrapping of microparticles by floppy lipid membranes

Lipid membranes form the barrier between the inside and outside of cells and many of their subcompartments. As such, they bind to a wide variety of nano- and micrometer sized objects and, in the presence of strong adhesive forces, strongly deform and envelop particles. This wrapping plays a key role in many healthy and disease-related processes. So far, little work has focused on the dynamics of wrapping. Here, using a model system of micron-sized colloidal particles and giant unilamellar lipid vesicles with tunable adhesive forces, we measure the velocity of the particle during wrapping as well as the forces exerted on it by the lipid membrane. Dissipation near the contact line appears to be the main factor determining the wrapping velocity and time to wrap an object

A correlative approach to segmenting phases and ferrite morphologies in transformation-induced plasticity steel using electron back-scattering diffraction and energy dispersive X-ray spectroscopy

Using a combination of electron back-scattering diffraction and energy dispersive X-ray spectroscopy data, a segmentation procedure was developed to comprehensively distinguish austenite, martensite, polygonal ferrite, ferrite in granular bainite and bainitic ferrite laths in a thermo-mechanically processed low-Si, high-Al transformation-induced plasticity steel. The efficacy of the ferrite morphologies segmentation procedure was verified by transmission electron microscopy. The variation in carbon content between the ferrite in granular bainite and bainitic ferrite laths was explained on the basis of carbon partitioning during their growth

Geometrical frustration of phase-separated domains in Coscinodiscus diatom frustules

Diatoms are single-celled organisms with a cell wall made of silica, called the frustule. Even though their elaborate patterns have fascinated scientists for years, little is known about the biological and physical mechanisms underlying their organization. In this work, we take a top-down approach and examine the micrometer-scale organization of diatoms from the Coscinodiscus family. We find two competing tendencies of organization, which appear to be controlled by distinct biological pathways. On one hand, micrometer-scale pores organize locally on a triangular lattice. On the other hand, lattice vectors tend to point globally toward a center of symmetry. This competition results in a frustrated triangular lattice, populated with geometrically necessary defects whose density increases near the center.ISSN:0027-8424ISSN:1091-649

Surface Passivation Method for the Super-repellence of Aqueous Macromolecular Condensates

Solutions of macromolecules can undergo liquid-liquid phase separation to form droplets with ultralow surface tension. Droplets with such low surface tension wet and spread over common surfaces such as test tubes and microscope slides, complicating in vitro experiments. The development of a universal super-repellent surface for macromolecular droplets has remained elusive because their ultralow surface tension requires low surface energies. Furthermore, the nonwetting of droplets containing proteins poses additional challenges because the surface must remain inert to a wide range of chemistries presented by the various amino acid side chains at the droplet surface. Here, we present a method to coat microscope slides with a thin transparent hydrogel that exhibits complete dewetting (contact angles θ ≈ 180°) and minimal pinning of phase-separated droplets in aqueous solution. The hydrogel is based on a swollen matrix of chemically cross-linked polyethylene glycol diacrylate of molecular weight 12 kDa (PEGDA), and can be prepared with basic chemistry laboratory equipment. The PEGDA hydrogel is a powerful tool for in vitro studies of weak interactions, dynamics, and the internal organization of phase-separated droplets in aqueous solutions.ISSN:0743-7463ISSN:1520-582

Surface Passivation Method for the Super-repellence of Aqueous Macromolecular Condensates

Solutions of macromolecules can undergo liquid–liquid phase separation to form droplets with ultralow surface tension. Droplets with such low surface tension wet and spread over common surfaces such as test tubes and microscope slides, complicating in vitro experiments. The development of a universal super-repellent surface for macromolecular droplets has remained elusive because their ultralow surface tension requires low surface energies. Furthermore, the nonwetting of droplets containing proteins poses additional challenges because the surface must remain inert to a wide range of chemistries presented by the various amino acid side chains at the droplet surface. Here, we present a method to coat microscope slides with a thin transparent hydrogel that exhibits complete dewetting (contact angles θ ≈ 180°) and minimal pinning of phase-separated droplets in aqueous solution. The hydrogel is based on a swollen matrix of chemically cross-linked polyethylene glycol diacrylate of molecular weight 12 kDa (PEGDA), and can be prepared with basic chemistry laboratory equipment. The PEGDA hydrogel is a powerful tool for in vitro studies of weak interactions, dynamics, and the internal organization of phase-separated droplets in aqueous solutions