Liposomes have established roles in drug delivery and cell membrane studies. Amongst other applications; they can also be used as analytical reagents, particularly in immunoassays. Liposomal immunoassays have potential advantages over alternatives; including sensitivity, speed, simplicity and relative reagent stability. The aim of these studies was to develop and evaluate novel examples of these assays. When liposomes entrapped the dye, Sulphorhodamine B, a shift in its maximum absorption wavelength compared to free dye was observed. This was attributed to dimerization of the dye at high concentrations. If the liposomes were disrupted, the released dye was diluted into the external buffer, and the dye's absorption spectrum reverted to that of free dye. After optimization of dye entrapment, immunoassays were developed using these liposomes. Albumin-coated liposomes were used in a model assay to measure serum albumin. This assay employed complement-mediated immunolysis, commonly used in liposomal immunoassays. The liposomes were lysed by anti-albumin and complement, and this could be competitively inhibited by serum albumin. To improve sensitivity, Fab' anti-albumin liposomes were prepared. These enabled measurement of urinary albumin by a complement-mediated immunoassay, but using a sandwich technique. Anti-albumin (intact) liposomes were shown to precipitate on gentle centrifugation after reaction with albumin. They were applied as a solid phase reagent in an heterogeneous immunoassay, using radioimmunoassay for urinary microalbumin as a model assay. Liposomes containing Sulphorhodamine B were also used in a more novel assay; for serum anticardiolipin antibodies. Cardiolipin-containing liposomes were prepared. These were lysable using magnesium ions. Anticardiolipin antibodies (IgG) were found to augment this lysis, enabling their estimation. Similar imprecision and acceptable correlation with a commercial enzyme-linked immunosorbent assay (ELISA) were obtained. The findings demonstrate Sulphorhodamine B release can be used as a marker in homogeneous colorimetric liposomal immunoassays; both in model assays and in potentially more useful clinical biochemistry applications.

Metal-polymer nanocomposites consisting of gold nanorods and temperature-responsive hydrogel nanoparticulates were investigated for use in externally-controlled drug delivery systems. Several different thermo-responsive hydrogels including poly(N-isopropyl acrylamide) (PNIPAAm) and poly(N-isopropryl acrylamide-co-acrylic acid) (P(NIPAAm-co-AA)) nanoparticles were synthesized for these nanocomposites using an aqueous dispersion polymerization method. In addition, nanoparticles of interpenetrating polymer networks (IPN) composed of poly(acrylamide) (PAAm) and poly(acrylic acid) (PAA) were synthesized using a water-in-oil emulsion polymerization. Temperature-responsive equilibrium swelling behavior of nanoparticles with varying crosslinking densities was characterized using dynamic light scattering. IPN systems exhibited a positive swelling response upon heating while PNIPAAm and copolymer systems collapsed upon increase in temperature above the transition point. Nanoparticles were characterized using scanning electron microscopy (SEM) and transmission electron microscopy (TEM) which demonstrated shape and morphology of polymer particles. Gold-polymer nanocomposites were formed by grafting gold nanorods to the surface of the polymer nanoparticles. Amine-functionalized gold nanorods were coupled to polymers using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) and N-hydroxysulfosuccinimide (Sulfo-NHS) to activate carboxyl groups on the surface of the polymer nanoparticles. TEM confirmed successful formation of the metal-polymer nanocomposites. Loading and release of a model therapeutic were done to assess the potential use of the polymer component of the nanocomposite for drug delivery. Fluorescein, a model for chemotherapeutics, was loaded into P(NIPAAm-co-AA) polymer nanoparticulates. Loading of the compound was shown to be a function of crosslinking density in the polymer network. Maximum loading was achieved using nanoparticles synthesized with a 10 mol% crosslinker feed ratio with entrapment efficiencies of 80.0 % and loading capacities of 12.0 %. Cytotoxicity studies were performed using a NIH/3T3 mouse fibroblast cell model. Cell viabilities in presence of P(NIPAAm-co-AA) nanoparticles were comparable to (not statistically different than) controls at concentrations up to 4 mg/ml. Similarly, gold-polymer composite concentrations up to 0.5 mg/ml caused limited cell death. / text

Submicron sized vesicles consisting of single and double chain non-ionic surfactant mixtures were prepared by simple dispersion of surfactant dissolved in aqueous medium, or alternatively, injecting the surfactant dissolved in organic solvent into the aqueous phase. Drug entrapment values were measured by using a fluorescent marker, 5,6- Carboxyfluorescein, and drug release characteristics were evaluated in biological media (serum and plasma) as a function of surfactant composition and in the presence or absence of cholesterol. Surface charge measurements, zeta-potential, as a function of pH, gel electrophoresis and immunoblotting (ELISA) were performed in order to measure the interaction of components of the biological fluid with the prepared vesicles. It was found that all vesicles carried a negative charge and rapidly bound plasma protein, which included albumin and immunoglobulin G, thus affecting the latency of the entrapped marker. Uptake and degradation of niosomes (non-ionic surfactant vesicles) in a living, unicellular, eukaryotic micro-organism was also investigated. It was found that the rate of release of contents depended on the composition of the vesicles and was a function of enzymatic degradation within these organisms rather than an intracellular PH effect of the digestive organelle. An identical protocol was carried out with the well- characterised liposome system and their inherent stabilities under a variety of conditions directly compared with niosomes.

To ensure the effective and safe use of nanomaterials for medical applications, the biocompatibility of the materials must be tested with particular relevance to the environment in which the material is placed. In nanoparticle-based drug delivery, it is crucial to evaluate a nanoparticle???s biocompatibility to ensure minimal cytotoxicity. Of several types of nanoparticles, peptide-based nanoparticles have emerged as promising systems for targeted cancer therapy. Yet, the biocompatibility of many of these peptides and their assembled particles has not been studied. This thesis, summarizes the original contribution on the effective and safe use of the particular self/co-assembling, amino acid pairing peptides and some of their DEGylated forms (modified versions) as carriers for anticancer drug delivery application. Therefore, the biocompatibility of the self-assembling, amino acid pairing (AAP) peptides AC8, its two DEGylated forms, as well as two related peptides, EAK16-II and EK8, is systematically investigated. The toxicity of these peptides and their complexes with pirarubicin was tested against the human adenocarcinoma lung cancer cell line, A549.The biocompatibility of the peptide-drug co-assembling complexes is assessed and the potential of these five peptides as carriers for the hydrophobic anticancer drug pirarubicin is demonstrated. For the first time experimental results on cytotoxicity, haemolytic activity, red blood cell (RBC) aggregation, complement activation and anaphylotoxin activation as an end result of complement activity for these five AAP peptides is reported. AC8, the amino end DEGylated AC8 (NP-I) and EK might be strong candidates for hydrophobic drug delivery considering their lack of toxicity and the fact that they are not recognized as a foreign molecule, inducing no considerable immune reactions. These results provide a basis for in vivo experiments and predict minimal in vitro toxicity of these peptides based delivery systems.

No description available.

No description available.

No description available.

This work introduces novel approaches to enhance targeting of pharmacotherapies to cornea, ciliary body, choroid, and posterior segment of the eye using microneedles as a drug delivery platform. The first part of the work determines the ability to deliver protein therapeutics into the cornea using coated microneedles to suppress corneal neovascularization in a rabbit model. The data show that highly targeted delivery of the anti-vascular endothelial growth factor protein therapeutic gave a better biological response of suppressing neovascularization with 11,900 times less dosage compared to topical administration. The second part of the research aims to develop novel formulations to target ciliary body and choroid via suprachoroidal delivery. The results show that a strongly non-Newtonian fluid can be used to slow down the spreading of the particles at the injection site up to 2 months. The results also show that a high molecular weight formulation with weakly non-Newtonian fluid can be used to reach 100% coverage of the choroidal surface with a single injection. The third part of the research aims to determine the biological response of targeting anti-glaucoma therapeutics to the ciliary body in a rabbit model. The results show we can achieve 500- to 1000-fold dose sparing by targeted delivery via supraciliary delivery. The fourth and last part of the research aims to develop novel emulsion droplets to target different locations within the eye using a gravity-mediated delivery technique via suprachoroidal space injection. The results show that we can deliver up to 73% of injected polymeric particles posterior to the equator of the eye. Overall this work demonstrates that microneedles have the capability to deliver pharmacotherapies to cornea, ciliary body, choroid, and posterior of the eye in a highly targeted manner and provide significant dose sparing in the rabbit model.

No description available.

No description available.