Plasmid DNA (pDNA) purification has been extensively investigated for various biological and biochemical applications such as transfection, polymerase chain reaction and DNA therapeutics. In the previous paper, we have described the synthesis, characterization and evaluation of microbeads (“Amikabeads”) derived from aminoglycoside amikacin for pDNA binding via anion-exchange chromatography. Here, we investigated the pDNA binding performance of conjugating Amikabeads with two highly specific DNA binding ligands via minor groove hydrophobic interaction. The pDNA maximum binding capacity of doxorubicin drug-conjugated Amikabeads (“doxo-beads”) was found out to be 429 μg pDNA/ mg of doxo-beads with a Langmuir constant of 8.21*10-4 L/mg, whereas the binding performance of berenil drug-conjugated "mikabeads (“berenil-beads”) was 142 μg pDNA/mg of berenil-beads with a adsorption constant of 4.71*10-5 L/mg. In addition, the desorption percentage of doxo-beads and berenil-beads was obtained as 52% and 41%, respectively. Our results indicate that by conjugating with highly specific DNA binding ligands, Amikabeads-drug complex enhances the pDNA binding performance and contains a promising potential for future applications in biotechnology field.

This research addresses the need for improvement in radiation sensors for applications of ionizing radiation such as radiotherapy. The current sensors involved are polymer gel dosimeters, MOSFETs, radio-chromic films, etc. Most of the sensors involved require expensive equipment's and processing facilities for readout. There is still a need to develop better sensors that can be clinically applied. There are numerous groups around the world trying to conceive a better dosimeter. One of the radiation sensors that was developed recently was based on fluorescence signal emitted from the sensor. To advance the field of radiation sensors, a visual indicator has been developed in-lab as a method of detect ionizing radiation. The intensity of change in color is directly dependent on the amount of incident ionizing radiation. An aqueous gold nanoparticle sensor can be used to accurately determine the incident amount of ionizing radiation1. A gold nanoparticle sensor has been developed in lab with the use of hexadecyltrimethylammonium bromide (C16TAB) as the templating molecule. In the presence of ionizing radiation, the colorless gold salt is reduced and templated, creating a dispersion within the fluid1. The formation of suspended nanoparticles leads to a color change that can be visually detected and accurately analyzed through the employment of a spectrometer. Unfortunately, the toxicity of C16TAB is high. It is expected the toxicity can be reduced by replacing C16TAB with an amino acid, as amino acids can act as templating molecules in the solution and many are naturally occuring2. The experiments included a screening of 20 natural amino acids and 12 unnatural amino acids with the gold salt solution in the presence of ionizing radiation. Stability and absorbance testing was conducted on the amino acid sensors. Additional screening of lead amino acid sensors at various concentrations of irradiation was conducted.

Genetic manipulation of human cell lines has widespread applications in biomedical research ranging from disease modeling to therapeutic development. Human cells are generally difficult to genetically engineer, but exogenous nucleic acids can be expressed by viral, chemical, or nonchemical means. Chemical transfections are simpler in practice than both viral and nonchemical delivery of genetic material, but often suffer from cytotoxicity and low efficiency. Novel aminoglycoside antibiotic-derived lipopolymers have been synthesized to mediate transgene delivery to human cells. These polymers are comprised of either paromomycin or apramycin crosslinked with glycerol diglycidylether and derivatized with stearoyl chloride in varying molar ratios. In this work, three previously identified target lipopolymers were screened against a library of human embryonic and induced pluripotent stem cell lines. Cells were transfected with a plasmid encoding green fluorescent protein (GFP) and expression was quantified with flow cytometry 48 hours after transfection. Transfection efficiency was evaluated between three distinct lipopolymers and four lipopolymer:DNA mass ratios. GFP expression was compared to that of cells transfected with commercially available chemical gene delivery reagent controls\u2014JetPEI, Lipofectamine, and Fugene\u2014at their recommended reagent:DNA ratios. Improved transgene expression in stem cell lines allows for improved research methods. Human stem cell-derived neurons that have been genetically manipulated to express phenotypic characteristics of aging can be utilized to model neurodegenerative diseases, elucidating information about these diseases that would be inaccessible in unmanipulated tissue.

Oxygen delivery is crucial for the development of healthy, functional tissue. Low tissue oxygenation, or hypoxia, is a characteristic that is common in many tumors. Hypoxia contributes to tumor malignancy and can reduce the success of chemotherapy and radiation treatment. There is a current need to noninvasively measure tumor oxygenation or pO2 in patients to determine a personalized treatment method. This project focuses on creating and characterizing nanoemulsions using a pO2 reporter molecule hexamethyldisiloxane (HMDSO) and its longer chain variants as well as assessing their cytotoxicity. We also explored creating multi-modal (MRI/Fluorescence) nanoemulsions.

The objective of the research presented here was to validate the use of kinetic models for the analysis of the dynamic behavior of a contrast agent in tumor tissue and evaluate the utility of such models in determining kinetic properties - in particular perfusion and molecular binding uptake associated with tissue hypoxia - of the imaged tissue, from concentration data acquired with dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) procedure. Data from two separate DCE-MRI experiments, performed in the past, using a standard contrast agent and a hypoxia-binding agent respectively, were analyzed. The results of the analysis demonstrated that the models used may provide novel characterization of the tumor tissue properties. Future research will work to further characterize the physical significance of the estimated parameters, particularly to provide quantitative oxygenation data for the imaged tissue.

The R-specific alcohol dehydrogenase (RADH or LVIS_0347) from Lactobacillus brevis LB19 was found to possess activity on several short chain aldehydes and ketones. This broad substrate specificity was previously uncharacterized. To demonstrate its relevance to the biofuels industry as well as its broader utility for chiral reductions, a detailed characterization was performed to further investigate the activity and function of RADH.

Increased investigation into the development of macromolecular fluorophores has resulted in the synthesis and discovery of several potential candidates. These include modified and polymeric based dendritic structures, hyperbranched polymers and linear polymers. Strong inherent blue photoluminescence has been recently described in linear polyamine polymers in the absence of any chemical modifications. Here we describe the screening of amine/polyamine compounds for inherent photoluminescence. Several compounds that exhibited strong inherent blue photoluminescence following excitation with UV light were identified. Furthermore we demonstrated successful synthesis of poly(amino ether) polymers as well as chemically cross-linked poly(amino ether) thermosets with the lead Pentaethylenehexamine which was found to have strong inherent blue photoluminescence. The polymers and thermosets were found to retain the photoluminescent properties of the original lead compound. The polymers and thermosets were investigated for their ability to sequester heavy metals from aqueous solutions. An increased decrease in initial photoluminescence was observed as the materials were incubated with increasing metal salt concentration as a result of metal binding sequestration. The poly(amino ether) polymers were found to have higher sensitivity for metal sequestration when compared to equivalent amount of linear 25 kDa polyethylenimine. The strong inherent blue photoluminescence and the ease of synthesis of the poly(amino ether) polymers and thermosets give these materials strong potential for future applications as sensors.

Abstract Molecular Engineering of Novel Polymeric Agents for Targeted Cancer Gene Therapy Dana Matthews Cancer gene cell therapy is a strategy that involves the administration of genes for correcting the effect of mutated cancer cells in order to induce tumor cell death. In particular, genes that encode for pro-apoptotic proteins can result in death of tumor cells. Prostate cancer is a very common cancer among males in America, and as highly destructive chemotherapy and radiation are generally the only treatments available once the cancer has metastasized, there is a need for the development of treatments that can specifically target and kill prostate cancer cells, while demonstrating low toxicity to other tissue. This experiment will attempt to create such a treatment through gene therapy techniques. The parallel synthesis and DNA binding affinity assay utilized in these experiments have produced a polymer that surpasses pEI-25, a gene delivery polymer standard, in both transfection efficacy and low cytotoxicity and trafficking of polyplexes in the cell, and finding methods to increase the transfection efficacy and specificity of polyplexes for PC3-PSMA cells.

Antibiotic resistant bacteria are a worldwide epidemic threatening human survival. Antimicrobial susceptibility tests (ASTs) are important for confirming susceptibility to empirical antibiotics and detecting resistance in bacterial isolates. Current ASTs are based on bacterial culturing, which take 2-14 days to complete depending on the microbial growth rate. Considering the high mortality and morbidity rates for most acute infections, such long time frames are clinically impractical and pose a huge risk to a patient's life. A faster AST will reduce morbidity and mortality rates, as well as help healthcare providers, administer narrow spectrum antibiotics at the earliest possible treatment stage.

In this dissertation, I developed a nonculture-based AST using an imaging and cell tracking technology. I track individual Escherichia coli O157:H7 (E. coli O157:H7) Uropathogenic Escherichia Coli (UPEC) cells, widely implicated in food-poisoning outbreaks and urinary tract infections respectively. Cells tethered to a surface are tracked on the nanometer scale, and phenotypic motion is correlated with bacterial metabolism. Antibiotic action significantly slows down motion of tethered bacterial cells, which is used to perform antibiotic susceptibility testing. Using this technology, the clinical minimum bactericidal concentration of an antibiotic against UPEC pathogens was calculated within 2 hours directly in urine samples as compared to 3 days using current gold standard tools.

Such technologies can make a tremendous impact to improve the efficacy and efficiency of infectious disease treatment. This has the potential to reduce the antibiotic mis-prescription steeply, which can drastically decrease the annual 2M+ hospitalizations and 23,000+ deaths caused due to antibiotic resistance bacteria along with saving billions of dollars to payers, patients, and hospitals.