Browsing by Author "Bickford, Lissett R."

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    Characterization of Conventional One-Step Sodium Thiosulfate Facilitated Gold Nanoparticle Synthesis
    Saverot, Scott-Eugene; Reese, Laura M.; Cimini, Daniela; Vikesland, Peter J.; Bickford, Lissett R. (SpringerOpen, 2015-05-28)
    Gold-gold sulfide nanoparticles are of interest for drug delivery, biomedical imaging, and photothermal therapy applications due to a facile synthesis method resulting in small particles with high near-infrared (NIR) absorption efficiency. Previous studies suggest that the NIR sensitivity of these nanoparticles was due to hexagonally shaped metal-coated dielectric nanoparticles that consist of a gold sulfide core and gold shell. Here, we illustrate that the conventional synthesis procedure results in the formation of polydisperse samples of icosahedral gold particles, gold nanoplates, and small gold spheres. Importantly, through compositional analysis, via UV/vis absorption spectrophotometry, transmission electron microscopy (TEM), and energy dispersive x-ray spectroscopy (EDS), we show that all of the nanoparticles exhibit identical face center cubic (FCC) gold crystalline structures, thus suggesting that sulfide is not present in the final fabricated nanoparticles. We show that icosahedrally shaped nanoparticles result in a blue-shifted absorbance, with a peak in the visible range. Alternatively, the nanoplate nanoparticles result in the characteristic NIR absorbance peak. Thus, we report that the NIR-contributing species in conventional gold-gold sulfide formulations are nanoplates that are comprised entirely of gold. Furthermore, polydisperse gold nanoparticle samples produced by the traditional one-step reduction of HAuCl4 by sodium thiosulfate show increased in vitro toxicity, compared to isolated and more homogeneous constituent samples. This result exemplifies the importance of developing monodisperse nanoparticle formulations that are well characterized in order to expedite the development of clinically beneficial nanomaterials.
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    Design and Fabrication of a Mask Projection Microstereolithography System for the Characterization and Processing of Novel Photopolymer Resins
    Lambert, Philip Michael (Virginia Tech, 2014-09-17)
    The goal of this work was to design and build a mask projection microstereolithography (MPμSL) 3D printing system to characterize, process, and quantify the performance of novel photopolymers. MPμSL is an Additive Manufacturing process that uses DLP technology to digitally pattern UV light and selectively cure entire layers of photopolymer resin and fabricate a three dimensional part. For the MPμSL system designed in this body of work, a process was defined to introduce novel photopolymers and characterize their performance. The characterization process first determines the curing characteristics of the photopolymer, namely the Critical Exposure (Ec) and Depth of Penetration (Dp). Performance of the photopolymer is identified via the fabrication of a benchmark test part, designed to determine the minimum feature size, XY plane accuracy, Z-axis minimum feature size, and Z-axis accuracy of each photopolymer with the system. The first characterized photopolymer was poly (propylene glycol) diacrylate, which was used to benchmark the designed MPμSL system. This included the achievable XY resolution (212 micrometers), minimum layer thickness (20 micrometers), vertical build rate (360 layers/hr), and maximum build volume (6x8x36mm3). This system benchmarking process revealed two areas of underperformance when compared to systems of similar design, which lead to the development of the first two research questions: (i) 'How does minimum feature size vary with exposure energy?' and (ii) 'How does Z-axis accuracy vary with increasing Tinuvin 400 concentration in the prepolymer?' The experiment for research question (i) revealed that achievable feature size decreases by 67% with a 420% increase in exposure energy. Introducing 0.25wt% of the photo-inhibitor Tinuvin 400 demonstrated depth of penetration reduction from 398.5 micrometers to 119.7 micrometers. This corresponds to a decrease in Z-axis error from 119% (no Tinuvin 400) to 9% Z-axis error (0.25% Tinuvin 400). Two novel photopolymers were introduced to the system and characterized. Research question (iii) asks 'What are the curing characteristics of Pluronic L-31 how does it perform in the MPμSL system?' while Research Question 4 similarly queries 'What are the curing characteristics of Phosphonium Ionic Liquid and how does it perform in the MPμSL system?' The Pluronic L-31 with 2wt% photo-initiator had an Ec of 17.2 mJ/cm2 and a Dp of 288.8 micrometers, with a minimum feature size of 57.3 ± 5.7 micrometers, with XY plane error of 6% and a Z-axis error of 83%. Phosphonium Ionic Liquid was mixed in various concentrations into two base polymers, Butyl Diacrylate (0% PIL and 10% PIL) and Poly Ethylene Dimethacrylate (5% PIL, 15% PIL, 25% PIL). Introducing PIL into either base polymer caused the Ec to increase in all samples, while there is no significant trend between increasing concentrations of IL in either PEGDMA or BDA and depth of penetration. Any trends previously identified between penetration depth and Z accuracy do not seem to extend from one resin to another. This means that overall, among all resins, depth of penetration is not an accurate way to predict the Z axis accuracy of a part. Furthermore, increasing concentrations of PIL caused increasing % error in both XY plane and Z-axis accuracy .
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    Development of Bacteria-Based Bio-Hybrid Delivery Systems: Fabrication, and Characterization of Chemotaxis and Quorum Sensing
    Sahari, Ali Akbar (Virginia Tech, 2014-10-09)
    Bio-hybrid approaches have recently provided a possible solution to address the challenge of on-board actuation, control and communication modules for micro/nanoscale cargo-carrying vehicles by integrating live prokaryotic or eukaryotic cells with synthetic objects. More specifically, because micro/nanoparticles are able to transport cargos efficiently and bacteria can play the role of targeted and selective delivery agents, a hybrid of these two can advance the current strategies for environmental monitoring, drug delivery and medical imaging. The main goal of this dissertation was to fabricate, assemble, and characterize different components of a mobile network of bacteria-based bio-hybrid systems for long-term applications in drug delivery and biosensing. First, a new library of bacteria-enabled delivery systems was developed by coupling live engineered bacteria with non-spherical particles and the transport of these bacteria-based systems was investigated in the absence and presence of chemical cues using microfluidic platforms. Next, a quorum-sensing (QS) based bacterial cell-cell communication network was characterized in a high-throughput manner in order to understand the coordinated behavior of the bacterial species ferrying the cargoes. Lastly, the QS behavior of a chemotactic population of the bacterial species in response to the endogenously produced signaling molecules was studied. The work presented in this dissertation lays the foundation for a well-characterized generation of bacteria-assisted cargo delivery devices with enhanced transport properties and capable of executing pre-programmed multi-agent coordinated tasks upon their arrival at the target site.
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    Innovating for Global Health through Community-Based Participatory Research: Design of Mechanical Suction Machines for Rural Health Clinics in Malawi
    Taylor, Ashley R. (Virginia Tech, 2016-09-21)
    Clinicians in low and middle-income countries (LMIC) face many challenges, including high patient-to-staff ratios, limited resources, and inconsistent access to electricity. This research aimed to improve health outcomes in LMIC through an enlightened understanding of challenges associated with healthcare technology. To understand LMIC barriers to acquiring, maintaining, and repairing medical equipment, a community-based participatory study was conducted at three clinical settings in southern Malawi. Thirty-six clinical staff participated in surveys and focus groups to provide information on medical device challenges. Results from the study emphasize the importance of community-based participatory innovation to improve global health. Many clinical staff expressed frustration regarding inability to prevent patient mortality attributed to equipment failure. Data from the community-based participatory study of medical technology conducted in Malawi revealed key insights for designing for low and middle income countries, and more specifically, for communities in southern Malawi. Specifically, partner communities identified mechanical suction machines as a top priority for design innovation. Working with technical and clinical staff in Malawian communities, a prototype mechanical suction machine was designed and constructed. This work suggests that engineers working in low and middle income countries face a unique sundry of design requirements that require an intimate understanding of the local community, including community leaders, community beliefs and values, and locally available resources. Technology innovation for global health should incorporate community expertise and assets, and health and technical education efforts should be developed to increase working knowledge of medical devices.
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    Microdevices for Investigating Pulsed Electric Fields-Mediated Therapies at Cellular and Tissue Level
    Bonakdar, Mohammad (Virginia Tech, 2016-06-29)
    Recent attempts to investigate living systems from a biophysical point of view has opened new windows for development of new diagnostic methods and therapies. Pulsed electric fields (PEFs) are a new class of therapies that take advantage of biophysical properties and have proven to be effective in drug delivery and treating several disorders including tumors. While animal models are commonly being used for development of new therapies, the high cost and complexity of these models along with the difficulties to control the electric field in the animal tissue are some of the obstacles toward the development of PEFs-based therapies. Microengineered models of organs or Organs-on-Chip have been recently introduced to overcome the hurdles of animal models and provide a flexible and cost-effective platform for early investigation of a variety of new therapies. In this study microfluidic platforms with integrated micro-sensors were designed, fabricated and employed to study the consequences of PEFs at the cellular level. These platforms were specifically used to study the effects of PEFs on the permeabilization of the blood-brain barrier for enhanced drug delivery to the brain. Different techniques such as fluorescent microscopy and electrical impedance spectroscopy were used to monitor the response of the cell monolayers under investigation. Irreversible electroporation is a new focal ablation therapy based on PEFs that has enabled ablation of tumors in a non-thermal, minimally invasive procedure. Despite promising achievements and treatment of more than 5500 human patients by this technique, real-time monitoring of the treatment progress in terms of the size of the ablated region is still needed. To address that necessity we have developed micro-sensor arrays that can be implemented on the ablation probe and give real-time feedback about the size of the ablated region by measuring the electrical impedance spectrum of the tissue.
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    Optimizing Emerging Healthcare Innovations in 3D Printing, Nanomedicine, and Imageable Biomaterials
    Reese, Laura Michelle (Virginia Tech, 2015-01-05)
    Emerging technologies in the healthcare industry encompass revolutionary devices or drugs that have the potential to change how healthcare will be practiced in the future. While there are several emerging healthcare technologies in the pipeline, a few key innovations are slated to be implemented clinically sooner based on their mass appeal and potential for healthcare breakthroughs. This thesis will focus on specific topics in the emerging technological fields of nanotechnology for photothermal cancer therapy, 3D printing for irreversible electroporation applications, and imageable biomaterials. While these general areas are receiving significant attention, we highlight the potential opportunities and limitations presented by our select efforts in these fields. First, in the realm of nanomedicine, we discuss the optimization and characterization of sodium thiosulfate facilitated gold nanoparticle synthesis. While many nanoparticles have been examined as agents for photothermal cancer therapy, we closely examine the structure and composition of these specific nanomaterials and discuss key findings that not only impact their future clinical use, but elucidate the importance of characterization prior to preclinical testing. Next, we examine the potential use of 3D printing to generate unprecedented multimodal medical devices for local pancreatic cancer therapy. This additive manufacturing technique offers exquisite design detail control, facilitating tools that would otherwise be difficult to fabricate by any other means. Lastly, in the field of imageable biomaterials, we demonstrate the development of composite catheters that can be visualized with near infrared imaging. This new biomaterial allows visualization with near infrared imaging, offering potentially new medical device opportunities that alleviate the use of ionizing radiation. This collective work emphasizes the need to thoroughly optimize and characterize emerging technologies prior to preclinical testing in order to facilitate rapid translation.
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    Rapid Stereomicroscopic Imaging of HER2 Overexpression in Ex Vivo Breast Tissue Using Topically Applied Silica-Based Gold Nanoshells
    Bickford, Lissett R.; Langsner, Robert J.; Chang, Joseph; Kennedy, Laura C.; Agollah, Germaine D.; Drezek, Rebekah (Hindawi, 2012-10-22)
    Tumor margin detection for patients undergoing breast conservation surgery primarily occurs postoperatively. Previously, we demonstrated that gold nanoshells rapidly enhance contrast of HER2 overexpression in ex vivo tissue sections. Our ultimate objective, however, is to discern HER2 overexpressing tissue from normal tissue in whole, nonsectioned, specimens to facilitate rapid diagnoses. Here, we use targeted nanoshells to quickly and effectively visualize HER2 receptor expression in intact ex vivo human breast tissue specimens. Punch biopsies of human breast tissue were analyzed after a brief 5-minute incubation with and without HER2-targeted silica-gold nanoshells using two-photon microscopy and stereomicroscopy. Labeling was subsequently verified using reflectance confocal microscopy, darkfield hyperspectral imaging, and immunohistochemistry to confirm levels of HER2 expression. Our results suggest that anti-HER2 nanoshells used in tandem with a near-infrared reflectance confocal microscope and a standard stereomicroscope may potentially be used to discern HER2-overexpressing cancerous tissue from normal tissue in near real time and offer a rapid supplement to current diagnostic techniques.
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    Single Cell Biomechanical Phenotyping using Microfluidics and Nanotechnology
    Babahosseini, Hesam (Virginia Tech, 2016-01-20)
    Cancer progression is accompanied with alterations in the cell biomechanical phenotype, including changes in cell structure, morphology, and responses to microenvironmental stress. These alterations result in an increased deformability of transformed cells and reduced resistance to mechanical stimuli, enabling motility and invasion. Therefore, single cell biomechanical properties could be served as a powerful label-free biomarker for effective characterization and early detection of single cancer cells. Advances and innovations in microsystems and nanotechnology have facilitated interrogation of the biomechanical properties of single cells to predict their tumorigenicity, metastatic potential, and health state. This dissertation utilized Atomic Force Microscopy (AFM) for the cell biomechanical phenotyping for cancer diagnosis and early detection, efficacy screening of potential chemotherapeutic agents, and also cancer stem-like/tumor initiating cells (CSC/TICs) characterization as the critical topics received intensive attention in the search for effective cancer treatment. Our findings demonstrated the capability of exogenous sphingosine to revert the aberrant biomechanics of aggressive cells and showed a unique, mechanically homogeneous, and extremely soft characteristic of CSC/TICs, suitable for their targeted isolation. To make full use of cell biomechanical cues, this dissertation also considered the application of nonlinear viscoelastic models such as Fractional Zener and Generalized Maxwell models for the naturally complex, heterogeneous, and nonlinear structure of living cells. The emerging need for a high-throughput clinically relevant alternative for evaluating biomechanics of individual cells led us to the development of a microfluidic system. Therefore, a high-throughput, label-free, automated microfluidic chip was developed to investigate the biophysical (biomechanical-bioelectrical) markers of normal and malignant cells. Most importantly, this dissertation also explored the biomechanical response of cells upon a dynamic loading instead of a typical transient stress. Notably, metastatic and non-metastatic cells subjected to a pulsed stress regimen exerted by AFM exhibited distinct biomechanical responses. While non-metastatic cells showed an increase in their resistance against deformation and resulted in strain-stiffening behavior, metastatic cells responded by losing their resistance and yielded slight strain-softening. Ultimately, a second generation microfluidic chip called an iterative mechanical characteristics (iMECH) analyzer consisting of a series of constriction channels for simulating the dynamic stress paradigm was developed which could reproduce the same stiffening/softening trends of non-metastatic and metastatic cells, respectively. Therefore, for the first time, the use of dynamic loading paradigm to evaluate cell biomechanical responses was used as a new signature to predict malignancy or normalcy at a single-cell level with a high (~95%) confidence level.
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    A Sustainable Engineering Solution for Paediatric Dehydration in Low-Resource Clinical Environments
    Taylor, Ashley R.; Turovskiy, Jeffrey; Drew, Benjamin; Muelenaer, Andre A.; Redican, Kerry J.; Kochersberger, Kevin B.; Bickford, Lissett R. (Engineers Without Borders Australia, 2016)
    Engineering efforts in low resource environments pose a unique set of challenges, requiring an in-depth understanding of local needs, comprehensive mapping of community resources, and extensive collaboration with local expertise. The importance of these principles is demonstrated in this paper by detailing the novel design and field demonstration of an affordable, locally manufactured intravenous fluid regulation device. Collaboration with clinical personnel in Uganda and Malawi guided device design. In-country physicians emphasised the need to regulate volume of intravenous (IV) fluid delivered to a paediatric patient without use of electricity. The proposed device regulates IV fluid delivery within ±20 mL of total prescribed dosage, providing a method of reducing fatalities caused by over-hydration in low resource environments; the feasibility of building the device from local resources was demonstrated by a field research team in Malawi. The device was successfully constructed entirely from local resources for a total cost of $46.21 (USD). Additionally, the device was demonstrated in rural clinics where 89 % of surveyed clinical staff reported that they would use the device to regulate IV fluid delivery. This paper emphasises the importance of collaborating with communities for community-based engineering solutions. Mapping community assets and collaborating with local expertise are crucial to success of engineering efforts. Long-term, community-based efforts are likely to sustainably improve health outcomes and strengthen economies of communities worldwide.
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    Transcriptional Interference Regulates the Evolutionary Development of Speech
    Mortlock, Douglas P.; Fang, Zhi-Ming; Chandler, Kelly J.; Hou, Yue; Bickford, Lissett R.; de Bock, Charles E.; Eapen, Valsamma; Clarke, Raymond A. (MDPI, 2022-07-04)
    The human capacity to speak is fundamental to our advanced intellectual, technological and social development. Yet so very little is known regarding the evolutionary genetics of speech or its relationship with the broader aspects of evolutionary development in primates. In this study, we describe a large family with evolutionary retrograde development of the larynx and wrist. The family presented with severe speech impairment and incremental retrograde elongations of the pisiform in the wrist that limited wrist rotation from 180° to 90° as in primitive primates. To our surprise, we found that a previously unknown primate-specific gene TOSPEAK had been disrupted in the family. TOSPEAK emerged de novo in an ancestor of extant primates across a 540 kb region of the genome with a pre-existing highly conserved long-range laryngeal enhancer for a neighbouring bone morphogenetic protein gene GDF6. We used transgenic mouse modelling to identify two additional GDF6 long-range enhancers within TOSPEAK that regulate GDF6 expression in the wrist. Disruption of TOSPEAK in the affected family blocked the transcription of TOSPEAK across the 3 GDF6 enhancers in association with a reduction in GDF6 expression and retrograde development of the larynx and wrist. Furthermore, we describe how TOSPEAK developed a human-specific promoter through the expansion of a penta-nucleotide direct repeat that first emerged de novo in the promoter of TOSPEAK in gibbon. This repeat subsequently expanded incrementally in higher hominids to form an overlapping series of Sp1/KLF transcription factor consensus binding sites in human that correlated with incremental increases in the promoter strength of TOSPEAK with human having the strongest promoter. Our research indicates a dual evolutionary role for the incremental increases in TOSPEAK transcriptional interference of GDF6 enhancers in the incremental evolutionary development of the wrist and larynx in hominids and the human capacity to speak and their retrogression with the reduction of TOSPEAK transcription in the affected family.