Previously we reported emulsion polymerization of propylene sulfide with Pluronic F127 as an emulsifier, yielding nanoparticles (NPs) in the 25 nm size range. Immunologically functional NPs were prepared by adding an antigen-Pluronic conjugate to the polymerization mixture ( Reddy , S. T. , et al. ( 2007 ) Nat. Biotechnol. 25, 1159 ). We sought a more flexible scheme for conjugation of antigens and other biomolecules to the NP surfaces that would allow for milder reaction conditions than achievable during the polymerization step. Here, we present the synthesis of such functionalizable NPs in the form of NPs that carry thiol-reactive groups, to which thiol-containing antigens (peptide or protein) or other biomolecules can be conjugated under mild conditions to yield immunofunctional NPs. The Pluronic-stabilized poly(propylene sulfide) (PPS) NPs with thiol-reactive pyridyl disulfide groups are prepared in two steps by (1) emulsion polymerization of propylene sulfide in the presence of a carboxylate-Pluronic and (2) reaction of the carboxylic acid groups on the NP surface with cysteamine pyridyl disulfide and a water-soluble carbodiimide reagent. We choose pyridyl disulfide groups to have a reduction-sensitive disulfide bond linking the antigen to the NP surface, allowing efficient release of antigen inside the cell in response to the reductive conditions within the endosome. The functionalizable NPs are characterized by proton NMR, dynamic light scattering (DLS), UV/vis spectroscopy, and transmission electron microscopy (TEM). Conjugation of small molecules and protein to the NP surface is presented.
During COVID19 and other viral pandemics, rapid generation of host and pathogen genomic data is critical to tracking infection and informing therapies. There is an urgent need for efficient approaches to this data generation at scale. We have developed a scalable, high throughput approach to generate high fidelity low pass whole genome and HLA sequencing, viral genomes, and representation of human transcriptome from single nasopharyngeal swabs of COVID19 patients.
The synthetic peptide RAD16-II has shown promise in tissue engineering and drug delivery. It has been studied as a vehicle for cell delivery and controlled release of IGF-1 to repair infarcted cardiac tissue, and as a scaffold to promote capillary formation for an in vitro model of angiogenesis. The structure of RAD16-II is hierarchical, with monomers forming long β-sheets that pair together to form filaments; filaments form bundles approximately 30-60 nm in diameter; branching networks of filament bundles form macroscopic gels. We investigate the mechanics of shearing between the two β-sheets constituting one filament, and between cohered filaments of RAD16-II. This shear loading is found in filament bundle bending or in tensile loading of fibers composed of partial-length filaments. Molecular dynamics simulations show that time to failure is a stochastic function of applied shear stress, and that for a given loading time behavior is elastic for sufficiently small shear loads. We propose a coarse-grained model based on Langevin dynamics that matches molecular dynamics results and facilities extending simulations in space and time. The model treats a filament as an elastic string of particles, each having potential energy that is a periodic function of its position relative to the neighboring filament. With insight from these simulations, we discuss strategies for strengthening RAD16-II and similar materials.
Hydrogels formed from self-assembling synthetic oligopeptides have been studied for almost 2 decades for use in tissue engineering and drug delivery. Although a great deal has been learned about the microstructure of these materials, there remain questions about how peptide filaments are ordered to form a gel. These unanswered questions leave a disconnect between our understanding of the observed nanoscale mechanical properties of peptide filaments and the macroscale properties of the gels they constitute. This study helps to bridge this gap by examining the role of filament length and interfilament crosslinks in determining bulk mechanical properties. Microindentation was used for mechanical characterization, and microstructure was observed using thin-section transmission electron microscopy images of gels embedded in resin. Results suggest a gel structure in which filaments are not densely bundled as previously suggested and confirm that crosslinking can be an effective strategy to increase the stiffness of self-assembling oligopeptide gels.
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