• Fabrication of Polystyrene Core-Silica Shell Nanoparticles for Scintillation Proximity Assay (SPA) Biosensors

      Aspinwall, Craig A.; Noviana, Eka; Heien, Michael L.; Pyun, Jeffrey (The University of Arizona., 2015)
      The development of analytical tools for investigating biological pathways on the molecular level has provided insight into diseases and disorders. However, many biological analytes such as glucose and inositol phosphate(s) lack the optical or electrochemical properties needed for detection, making molecular sensing challenging. Scintillation proximity assay (SPA) does not require analytes to possess such properties. SPA uses radioisotopes to monitor the binding of analytes to SPA beads. The beads contain scintillants that emit light when the radiolabeled analytes are in close proximity. This technique is rapid, sensitive and separation-free. Conventional SPA beads, however, are large relative to the cells and made of hydrophobic organic polymers that tend to aggregate or inorganic crystals that sediment rapidly in aqueous solution, thus limiting SPA applications. To overcome these problems, polystyrene core-silica shell nanoparticles (NPs) doped with pTP and dimethyl POPOP were fabricated to produce scintillation NPs that emit photons in the blue region of visible light. The developed scintillation particles are approximately 250 nm in diameter (i.e. 200 nm of core diameter and 10-30 nm of shell thickness), responsive to β-decay from tritium (³H) and have sufficient stability in the aqueous media. DNA hybridization-based SPA was performed to determine whether the scintillation NPs could be utilized for SPA applications. A 30-mer oligonucleotide was immobilized on the polystyrene core-silica shell NPs to give approximately 7.6 x 10³ oligonucleotide molecules per NP and ³H-labeled complementary strand was annealed to the immobilized strand. At the saturation point, increases in scintillation signal due to oligonucleotide binding to the NPs were about 9 fold compared to the control experiments in which no specific binding occurred, demonstrating that the scintillation NPs can be utilized for SPA. Along with the improved physical properties including smaller size and better stability in the aqueous system, the developed scintillation NPs could be potentially useful as biosensors in cellular studies.
    • Nanoparticles Of PLGA With Encapsulated Insulin For Oral Controlled Release For Diabetes Treatment

      Guzman, Roberto; Abduljawad, Marwan; Guzman, Roberto; Gervasio, Dominic; Sorooshian, Armin (The University of Arizona., 2015)
      Insulin, a relatively low molecular weight protein has been used for decades in the treatment of diabetes; it has well-defined properties and delivery requirements. Due to the current increase of diabetes in the world improved insulin delivery systems could significantly influence the treatment of diabetes and the quality of life of the affected people. The main objective of this work was to encapsulate insulin in polymer nanoparticles of Poly (DL-Lactic-Co-Glycolic Acid) (PLGA) and poly vinyl alcohol (PVA). Preliminary results of these functional therapeutic nanoparticles prepared with PVA and PLGA by using a double emulsion method (water/oil/water) were obtained in terms of encapsulation efficiency and effective insulin release from the nanoparticles. Assessing the bioactivity of insulin once encapsulated and released is not trivial, thus an indirect protein assay was developed to effectively and easily assess the activity of proteins going through these processes. Trypsin, a proteolitic enzyme was used as model protein to investigate the biological activity of encapsulated and released biomolecules. The activity of trypsin towards a synthetic substrate, DL-BAPNA was used to measure the enzyme kinetics and activity before encapsulation, while encapsulated and after the enzyme was released from the nanoparticles. Results show that the enzyme maintained substantial activity while encapsulated and after its release. It is anticipated that the biological activity after being released from the nanoparticles will remain biologically active, however, biological assays remain to be performed to corroborate this argument. In addition to release experiments with trypsin and insulin, other proteins were also studied. In all cases the release form the nanoparticles at 37 °C exhibited a three stage release process, The release process will be modeled according to developed mathematical models that consider initial burst of molecules, degradation of polymer and diffusion of molecules from the nanoparticles.
    • Synthesis and Characterization of Ferromagnetic Polymer-Coated Cobalt Nanoparticles in Multi-Gram Quantities

      Pyun, Jeffrey; Rasmussen, Sarah Grace; Pyun, Jeffrey; Christie, Hamish; Loy, Douglas (The University of Arizona., 2009)
      Ferromagnetic cobalt nanoparticles (CoNPs) are of interest due to their inherent dipolar properties which enable one-dimensional (1-D) nanoparticle self-assembly. As their magnetic properties change drastically with their size, the ability to selectively synthesize monodisperse metallic nanoparticles of varying diameters remains a crucial challenge. Although there have been extensive studies performed on various metallic nanoparticles yielding superparamagnetic materials (such as Fe3O4, Fe2O3, Co metals), research concerning the synthesis of ferromagnetic materials has only recently resurged within the last 20 years.In this work, methods for the synthesis of ferromagnetic cobalt nanoparticles on multi-gram scales were investigated. A one-pot synthetic method which produced up to 4 grams of cobalt nanoparticles per reaction was developed, and it was also found that this reaction had a direct correlation with particle size and reaction temperature, allowing for the large-scale synthesis of polystyrene-coated cobalt nanoparticles of pre-selected diameters.