Long chain lipids facilitate insertion of large nanoparticles into membranes of small unilamellar vesicles

Insertion of hydrophobic nanoparticles into phospholipid bilayers is limited to small particles that can incorporate into the hydrophobic membrane core in between the two lipid leaflets. Incorporation of nanoparticles above this size limit requires development of challenging surface engineering methodologies. In principle, increasing membrane thickness should facilitate incorporation of larger nanoparticles. Here we explore the effect of very long phospholipids (C24:1) in small unilamellar vesicles, on the membrane insertion efficiency of nanoparticles in the range of 5-13 nm in diameter. To this end, we improved an existing vesicle preparation protocol and utilized cryogenic electron microscopy imaging to examine the mode of interaction and to evaluate the membrane insertion efficiency of membrane-inserted nanoparticles.

Giant Vesicles and Their Use in Assays for Assessing Membrane Phase State, Curvature, Mechanics, and Electrical Properties

Giant unilamellar vesicles represent a promising and extremely useful model biomembrane system for systematic measurements of mechanical, thermodynamic, electrical, and rheological properties of lipid bilayers as a function of membrane composition, surrounding media, and temperature. The most important advantage of giant vesicles over other model membrane systems is that the membrane responses to external factors such as ions, (macro)molecules, hydrodynamic flows, or electromagnetic fields can be directly observed under the microscope. Here, we briefly review approaches for giant vesicle preparation and describe several assays used for deducing the membrane phase state and measuring a number of material properties, with further emphasis on membrane reshaping and curvature.

Immunization withSalmonella entericaSerovar Typhimurium-Derived Outer Membrane Vesicles Delivering the Pneumococcal Protein PspA Confers Protection against Challenge withStreptococcus pneumoniae

ABSTRACTGram-negative bacteria produce outer membrane vesicles (OMVs) that serve a variety of functions related to survival and pathogenicity. Periplasmic and outer membrane proteins are naturally captured during vesicle formation. This property has been exploited as a method to derive immunogenic vesicle preparations for use as vaccines. In this work, we constructed aSalmonella entericaserovar Typhimurium strain that synthesized a derivative of the pneumococcal protein PspA engineered to be secreted into the periplasmic space. Vesicles isolated from this strain contained PspA in the lumen. Mice intranasally immunized with the vesicle preparation developed serum antibody responses against vesicle components that included PspA andSalmonella-derived lipopolysaccharide and outer membrane proteins, while no detectable responses developed in mice immunized with an equivalent dose of purified PspA. Mucosal IgA responses developed against theSalmonellacomponents, while the response to PspA was less apparent in most mice. Mice immunized with the vesicle preparation were completely protected against a 10× 50% lethal dose (LD50) challenge ofStreptococcus pneumoniaeand significantly protected against a 200× LD50challenge, while control mice immunized with purified PspA or empty vesicles were not protected. These results establish that vesicles can be used to mucosally deliver an antigen from a Gram-positive organism and induce a protective immune response.