The capability to resolve the contributions from spectroscopically overlapping fluorophores has enabled significant breakthroughs in cellular imaging. However, commercial microscopes for this purpose use analog light detection with least squares curve-fitting analysis and improvements in sensitivity are needed. To this end, a microscope has been constructed with high throughput and single-photon detection capability. The fluorescence is separated through use of a prism spectrometer or a series of dichroic mirrors into four spectral bands and detected using four single-photon avalanche diode (SPAD) detectors, which provide high-quantum efficiency in the red spectral region. The detectors are connected to a time-correlated single photon counting module to provide sub-nanosecond temporal resolution for distinguishing fluorophores with different fluorescence lifetimes. Maximum-likelihood (ML) methods have been developed for analyzing the temporally and spectrally resolved photon count data from the SPADs to find the contributions from different fluorescent species and from background. Commercially available SPADs exhibit a count-rate dependent time shift in the impulse response function, and hence the instrument incorporates custom modified SPADs with improved timing stability. Nevertheless, there is still some time shift, and hence the ML-analysis has been extended to include this as an adjustable parameter for each individual SPAD. Monte Carlo simulations have also been developed to enable studies of the number of photons needed to resolve specific fluorophores.

Most biological experiments are performed on an ensemble of cells under the assumption that all cells are identical. However, recent evidence from single cells studies reveals that this assumption is incorrect. Individual cells within the same generation may differ dramatically, and these differences have important consequences for the health and function of the entire living body. I have used Scanning Ion Conductance Microscopy (SICM) for imaging and analysis of topographical change of single cell membrane, which is difficult to be revealed by optical microscopes. Morphological change in the fixed and live HeLa cell membrane during endocytosis of conjugated polymer nanoparticles was studied. Results demonstrated SICM is a powerful tool to study the interaction between nanoparticle and cell membrane during internalization of nanoparticles through the membrane. This research can improve our fundamental understanding of cellular behavior and will be helpful for drug delivery applications. Based on conventional SICM, we have developed a novel method to simultaneous map the topography and potential distributions of the single living cells membranes. At the first step, multifunctional nanopipettes (nanopore/nanoelectrode) have been fabricated and characterized. To demonstrate the potential sensing capability and understand the mechanism, I measured the ionic current and local electric potential change during translocation of 40 nm charged gold nanoparticles. Our results reveal the capability of the multifunctional probe for the highly sensitive detection of the ionic current and local electrical potential changes during the translocation of the charged entity through the nanopore. From the potential change, we revealed the dynamic assembly of GNPs before entering the nanopore. The experimental results are also nicely explained by the finite element method based numerical simulation results. At the second step, I have measured the surface potential of living cell membrane at selected locations. Very recently, I have obtained results to show that we can map the extracellular membrane potential distribution of the complicated living cell membrane with sub-micron spatial resolution.This new imaging technique can help biologist to explore the extracellular potential distribution of varieties of cells quantitatively.These studies will have impacts on several biomedical applications such as regenerative repair and cancer treatment.

Nanomaterials such as graphene oxide and carbon nanotubes, have demonstrated excellent properties for membrane desalination, including decrease of maintenance, increase of flux rate, simple solution casting, and impressive chemical inertness. Here, two projects are studied to investigate nanocarbon based membrane desalination. The first project is to prepare hybrid membranes with amyloid fibrils intercalated with graphene oxide sheets. The addition of protein amyloid fibrils expands the interlayer spacing between graphene oxide nanosheets and introduces additional functional groups in the diffusion pathways, resulting in increase of flux rate and rejection rate for the organic dyes. Amyloid fibrils also provide structural assistance to the hybrid membrane, which supresses cracking and instability of graphene oxide sheets. The second project is to fabricate polymer nanocomposite membranes with carbon nanotubes encapsulated by polymerized surfactants. The designed polymerizable surfactant forms lyotropic liquid crystalline mesophases in an aqueous medium with hexagonal packing of cylindrical micelles. The adsorption of surfactants on the surface of carbon nanotubes allows a stable dispersion of carbon nanotubes encapsulated in the cylindrical micelles, resulting in the ordered structure. After photo-polymerization, the composite membranes display enhanced dye rejection. Both projects have shown promising ways to improve membrane filtration by using nanomaterials.

<p>In this thesis, the ripple formation at the interface of SiO2 and Si were studied in a systematic fashion by irradiating the SiO2-Si samples with ultrashort laser pulses under a broad variety of experimental conditions. They consist of di↵erent irradiating laser wavelengths, incident laser energies, translation speeds, translation directions, spot sizes of the laser beam, as well as oxide thicknesses. The ripples produced by laser irradiation are examined using various microscopy techniques in order to characterize their surface morphology, detailed structures, crystalline properties, and so on. For the experiments carried out at ! = 800 nm, the ripples formed on the SiO2-Si sample with an oxide thickness of 216 nm were first observed under optical microscopy and SEM. After removing the oxide layer with HF solution, the surface features of the ripples on the Si substrate were investigated using SEM and AFM techniques. Subsequently, by means of TEM and EDX analysis, the material composition and crystallinity of the ripples were determined. It is concluded that the ripples are composed of nano-crystalline silicon. In addition to the 216 nm oxide thickness, other oxide samples with di↵erent oxide thicknesses, such as 24, 112, 117, 158 and 1013 nm, were also processed under laser irradiation. The ripple formation as a function of the laser energy, the translation direction and the spot size is discussed in detail. Furthermore, the ripples created at the SiO2-Si interface are compared with</p> <p>the LIPSS created on pure silicon samples that were processed under similar laser irradiation conditions. The spatial periodicities of the ripples were evaluated to be in the range of between 510 nm and 700 nm, which vary with the oxide thickness and other laser parameters. For the experiments using the ! = 400 nm laser pulses, it is found that ripples can also be formed at the SiO2-Si interface, which have spatial periodicities in the range of between 310 nm and 350 nm depending on the oxide thickness. The ripple formation at this 400 nm wavelength as a function of the laser energy, the translation speed, and translation direction is considered as well. For the case of ! = 400 nm irradiation, a comparison is also made between the interface ripples on the SiO2-Si samples and the LIPSS on a pure Si sample. Through FIB-TEM and EDX analysis, it confirmed that the ripples were produced in the substrate while the oxide layer maintained its structural integrity. In addition, the ripples are composed of nano-crystalline silicon whose crystallite sizes are on the order of a few nanometers. Apart from irradiating oxide samples with femtosecond laser pulses, which applies to the two cases of ! = 800 and 400 nm mentioned above, oxide samples with an oxide thickness of 112 nm were irradiated with picosecond laser pulses at ! = 800 nm whose pulse durations are 1 ps and 5 ps, respectively. However, no regular ripples can be produced at the SiO2-Si interface while maintaining the complete integrity of the oxide layer.</p> / Master of Applied Science (MASc)

<p>The class of meteorites called carbonaceous chondrites are examples of material from the solar system which have been relatively unchanged from the time of their initial formation. We investigate the carbonaceous chondrite subclasses CI, CM, CR, CV, and CO, which contain high levels of water and organic material, including amino acids. These subclasses span petrologic types 1 through 3, indicating the degree of internal chemistry undergone by the meteoritic parent body. The goal of this thesis is two-fold: to obtain a comprehensive view of amino acid abundances and relative frequencies in carbonaceous chondrites, and to recreate these patterns via thermodynamic computational models.</p> <p>We collate available amino acid abundance data for a variety of meteorites to identify patterns in total abundance and relative frequencies. We consider only a set of 20 proteinogenic alpha-amino acids created via a specific chemical pathway called Strecker synthesis. We plot abundances of individual amino acids for each subclass, as well as total abundances across all subclasses. We see a predominance in abundance and variety of amino acids in the CM and CR subclasses, which contain concentrations of amino acids greater by several orders of magnitude than other carbonaceous subclasses. These subclasses correspond to an aqueous alteration temperature range of 200 deg. C to 400 deg. C. Within the CM2 and CR2 meteorites, we identify trends in the relative frequencies of amino acids in preparation for computational modeling.</p> <p>Now having a baseline observed amino acid abundance plot, we recreate both the total amino acid abundance pattern as well as the relative frequency of amino acids within the CM2 chondrite subclass using computational models. We use thermodynamic theory of Gibbs free energies to calculate the output of amino acids in a meteoritic parent body assuming chemical equilibrium and some set of initial concentrations of organic material. Our model recreates abundance patterns in the temperature range 200 deg. C to 400 deg. C, ~10⁵ parts-per billion (ppb), and the temperature range 400 deg. C to 500 deg. C, ~10² ppb. Our model does not fit well between temperatures of 150 deg. C to 200 deg. C. Our current model assumes a uniform composition of initial chemical reactants; likely an inhomogeneous composition would be a more accurate physical representation of a parent body. In addition, we match relative frequencies to observed frequencies for each amino acid in the CM2 subclass to well within an order of magnitude.</p> / Master of Science (MSc)

In this work, we design a linear, two step implicit finite difference method to approximate the solutions of a biological system that describes the interaction between a microbial colony and a surrounding substrate. Three separate models are analyzed, all of which can be described as systems of partial differential equations (PDE)s with nonlinear diffusion and reaction, where the biological colony grows and decays based on the substrate bioavailability. The systems under investigation are all complex models describing the dynamics of biological films. In view of the difficulties to calculate analytical solutions of the models, we design here a numerical technique to consistently approximate the system evolution dynamics, guaranteeing that nonnegative initial conditions will evolve uniquely into new, nonnegative approximations. This property of our technique is established using the theory of M-matrices, which are nonsingular matrices where all the entries of their inverses are positive numbers. We provide numerical simulations to evince the preservation of the nonnegative character of solutions under homogeneous Dirichlet and Neumann boundary conditions. The computational results suggest that the method proposed in this work is stable, and that it also preserves the bounded character of the discrete solutions.

Cell mass is an important indicator of cell health and status. A diverse set of techniques have been developed to precisely measure the masses of single cells, with varying degrees of technical complexity and throughput. Here, the development of a non-invasive, label-free optical technique, termed Live-Cell Interferometry (LCI), is described. Several applications are presented, including an evaluation of LCI’s utility for assessing drug response heterogeneity in patient-derived melanoma lines and the measurement of CD3+ T cell kinetics during hematopoietic stem cell transplantation. The characterization of mast cells during degranulation, the measurement of viral reactivation kinetics in Kaposi’s Sarcoma, and drug response studies in patient-derived xenograft models of triple-negative breast cancer are also discussed. Taken together, data from these studies highlight LCI’s versatility as a tool for clinical, translational, and basic research applications.

Improvement in the in vitro oxidative folding of disulfide-containing proteins, such as extracellular and pharmaceutically important proteins, is required. Traditional folding methods using small molecule aliphatic thiol and disulfide, such as glutathione (GSH) and glutathione disulfide (GSSG) are slow and low yielding. Small molecule aromatic thiols and disulfides show great potentiality because aromatic thiols have low pKa values, close to the thiol pKa of protein disulfide isomerase (PDI), higher nucleophilicity and good leaving group ability. Our studies showed that thiols with a positively charged group, quaternary ammonium salts (QAS), are better than thiols with negatively charged groups such as phosphonic acid and sulfonic acid for the folding of bovine pancreatic trypsin inhibitor (BPTI). An enhanced folding rate of BPTI was observed when the protein was folded with a redox buffer composed of a QAS thiol and its corresponding disulfide. Quaternary ammonium salt (QAS) thiols and their corresponding disulfides with longer alkyl side chains were synthesized. These QAS thiols and their corresponding disulfides are promising small molecule thiols and disulfides to fold reduced BPTI efficiently because these thiols are more hydrophobic and can enter the core of the protein. Conformational changes of disulfide-containing proteins during oxidative folding influence the folding pathway greatly. We performed the folding of BPTI using targeted molecular dynamics (TMD) simulation and investigated conformational changes along with the folding pathway. Applying a bias force to all atoms versus to only alpha carbons and the sulfur of cysteines showed different folding pathways. The formation of kinetic traps N' and N* was not observed during our simulation applying a bias force to all atoms of the starting structure. The final native conformation was obtained once the correct antiparallel β-sheets and subsequent Cys14-Cys38 distance were decreased to a bond distance level. When bias force was applied to only alpha carbons and the sulfur of cysteines, the distance between Cys14-Cys38 increased and decreased multiple times, a structure similar to the confirmation of N*, NSH were formed and native protein was ultimately obtained. We concluded that there could be multiple pathways of conformational folding which influence oxidative folding.

Dr. Kwan Hsu was neither a superstar nor a celebrity. Her name does not come up in conversations about important contributors to her field of biophysics nor is she instantly recognizable for her contributions to Portland State University's international program or the state of Oregon's business ties with China. Yet she was a contributor, a cog-in-the-wheel, at the very least, in all of these areas and more. She was a peripheral member of a well-known Chinese family, but few in the United States know of or perhaps have interest in, but otherwise, she had no great connections or family ties to generate interest in her story. How does one process a collection for a woman who does not meet the traditional criteria for excellence or success or public interest for an archive? Where is the value to the larger historical narrative of our time in preserving the memories of someone who was non-remarkable, or, conversely, someone who may be even too unique to contribute to that greater narrative? These are the questions I wrestled with when I first came to this collection. As my research progressed, I realized that I faced more questions, and that to come to any understanding that might answer them, I was going to have to research the history of archives and archival processes. Science, the Cold War, Communist China, women, the immigrant experience, all of these issues became part of my thesis, however shallowly I was able to investigate them. Questions of identity and historiography, of power and discourse were explored. In the end, what I found was that a collection that on the outside looked unimpressive and unenlightening, could indeed be very valuable, and provide insight into any number of areas of current interest in historical research. This is that story.

Optical scattering techniques are suitable probes for studying weak disordered refractive index media such as biological cells and tissues. Several brain diseases accompany the nano-to-submicron scales’ structural alterations of the basic building blocks of cells/tissues in the brain and skin fibroblasts. For example, several molecular modifications such as DNA methylation, and histone degradation occur in cells earlier than morphological changes detectable at a microscopic level. These alterations also change the refractive index structures of the cells/tissues at the nano-to-submicron scales. Unfortunately, traditional methods do not allow the detection of these alterations in the early stages of diseases. Recent developments in mesoscopic optical physics-based techniques can probe these alterations. Particularly, mesoscopic light transport and localization approaches enable the measurements and quantifications of the degree of structural alterations in the cells/tissues and unprecedented information on progressive brain diseases. This dissertation provides a detailed study of the structural changes at nano-to-submicron levels in human brain cells/tissues and human skin fibroblasts in two major neurodegenerative diseases, Alzheimer’s disease (AD) and Parkinson's disease (PD), using dual spectroscopic imaging techniques, namely partial wave spectroscopy (PWS) for light transport and inverse participation ratio (IPR) for weak light localization. In particular, a nanoscale-sensitive advanced PWS technique is used to quantify the structural alterations in cells/tissues. Further, the IPR technique is used to quantify molecular-specific mass density alterations within cells using their light localization properties via confocal imaging. These dual optical scattering techniques were utilized to measure the degree of structural disorders, termed ‘disorder strength’, by distinguishing the diseased cells/tissues from normal ones in the human brain and human skin fibroblasts due to neurodegenerative diseases. Our results show that the degree of structural disorder (����) increases in the affected cells and tissues relative to the normal, both at the cellular/tissue level and in the DNA molecular mass density structural levels. The results of the studies strongly reveal that the degree of structural disorder strength (����) is an effective biomarker/numerical indicator for brain disease diagnostics.