Integration of virus-like particle macromolecular bioreceptors in electrochemical biosensors

Rapid, sensitive and selective detection of chemical hazards and biological pathogens has shown growing importance in the fields of homeland security, public safety and personal health. In the past two decades, efforts have been focusing on performing point-of-care chemical and biological detections using miniaturized biosensors. These sensors convert target molecule binding events into measurable electrical signals for quantifying target molecule concentration. However, the low receptor density and the use of complex surface chemistry in receptors immobilization on transducers are common bottlenecks in the current biosensor development, adding to the cost, complexity and time. This dissertation presents the development of selective macromolecular Tobacco mosaic virus-like particle (TMV VLP) biosensing receptor, and the microsystem integration of VLPs in microfabricated electrochemical biosensors for rapid and performance-enhanced chemical and biological sensing. Two constructs of VLPs carrying different receptor peptides targeting at 2,4,6-trinitrotoluene (TNT) explosive or anti-FLAG antibody are successfully bioengineered. The VLP-based TNT electrochemical sensor utilizes unique diffusion modulation method enabled by biological binding between target TNT and receptor VLP. The method avoids the influence from any interfering species and environmental background signals, making it extremely suitable for directly quantifying the TNT level in a sample. It is also a rapid method that does not need any sensor surface functionalization process. For antibody sensing, the VLPs carrying both antibody binding peptides and cysteine residues are assembled onto the gold electrodes of an impedance microsensor. With two-phase immunoassays, the VLP-based impedance sensor is able to quantify antibody concentrations down to 9.1 ng/mL. A capillary microfluidics and impedance sensor integrated microsystem is developed to further accelerate the process of VLP assembly on sensors and improve the sensitivity. Open channel capillary micropumps and stop-valves facilitate localized and evaporation-assisted VLP assembly on sensor electrodes within 6 minutes. The VLP-functionalized impedance sensor is capable of label-free sensing of antibodies with the detection limit of 8.8 ng/mL within 5 minutes after sensor functionalization, demonstrating great potential of VLP-based sensors for rapid and on-demand chemical and biological sensing.

Surface Insulation Resistance Degradation and Electrochemical Migration on Printed Circuit Boards

Widespread adoption of lead-free materials and processing for printed circuit board (PCB) assembly has raised reliability concerns regarding surface insulation resistance (SIR) degradation and electrochemical migration (ECM). As PCB conductor spacings decrease, electronic products become more susceptible to these failures mechanisms, especially in the presence of surface contamination and flux residues which might remain after no-clean processing. Moreover, the probability of failure due to SIR degradation and ECM is affected by the interaction between physical factors (such as temperature, relative humidity, electric field) and chemical factors (such as solder alloy, substrate material, no-clean processing). Current industry standards for assessing SIR reliability are designed to serve as short-term qualification tests, typically lasting 72 to 168 hours, and do not provide a prediction of reliability in long-term applications. The risk of electrochemical migration with lead-free assemblies has not been adequately investigated. Furthermore, the mechanism of electrochemical migration is not completely understood. For example, the role of path formation has not been discussed in previous studies. Another issue is that there are very few studies on development of rapid assessment methodologies for characterizing materials such as solder flux with respect to their potential for promoting ECM. In this dissertation, the following research accomplishments are described: 1). Long-term temp-humidity-bias (THB) testing over 8,000 hours assessing the reliability of printed circuit boards processed with a variety of lead-free solder pastes, solder pad finishes, and substrates. 2). Identification of silver migration from Sn3.5Ag and Sn3.0Ag0.5Cu lead-free solder, which is a completely new finding compared with previous research. 3). Established the role of path formation as a step in the ECM process, and provided clarification of the sequence of individual steps in the mechanism of ECM: path formation, electrodeposition, ion transport, electrodeposition, and filament formation. 4). Developed appropriate accelerated testing conditions for assessing the no-clean processed PCBs' susceptibility to ECM: a). Conductor spacings in test structures should be reduced in order to reflect the trend of higher density electronics and the effect of path formation, independent of electric field, on the time-to-failure. b). THB testing temperatures should be modified according to the material present on the PCB, since testing at 85oC can cause the evaporation of weak organic acids (WOAs) in the flux residues, leading one to underestimate the risk of ECM. 5). Correlated temp-humidity-bias testing with ion chromatography analysis and potentiostat measurement to develop an efficient and effective assessment methodology to characterize the effect of no-clean processing on ECM.

Advanced Adhesion Strength Testing Methods of Thin Film Multilayers in Electronic Packaging Systems

With the continued miniaturization and increasing performance of electronic devices, new technical challenges have arisen. One such issue is delamination occurring at critical interfaces inside the device. This major reliability issue can occur during the manufacturing process or during normal use of the device. Proper evaluation of the adhesion strength of critical interfaces early in the product development cycle can help reduce reliability issues and time-to-market of the product. However, conventional adhesion strength testing is inherently limited in the face of package miniaturization, which brings about further technical challenges to quantify design integrity and reliability. Although there are many different interfaces in today's advanced electronic packages, they can be generalized into two main categories: 1) rigid to rigid connections with a thin flexible polymeric layer in between, or 2) a thin film membrane on a rigid structure. Knowing that every technique has its own advantages and disadvantages, multiple testing methods must be enhanced and developed to be able to accommodate all the interfaces encountered for emerging electronic packaging technologies. For evaluating the adhesion strength of high adhesion strength interfaces in thin multilayer structures a novel adhesion test configuration called “single cantilever adhesion test (SCAT)” is proposed and implemented for an epoxy molding compound (EMC) and photo solder resist (PSR) interface. The test method is then shown to be capable of comparing and selecting the stronger of two potential EMC/PSR material sets. Additionally, a theoretical approach for establishing the applicable testing domain for a four-point bending test method was presented. For evaluating polymeric films on rigid substrates, major testing challenges are encountered for reducing testing scatter and for factoring in the potentially degrading effect of environmental conditioning on the material properties of the film. An advanced blister test with predefined area test method was developed that considers an elasto-plastic analytical solution and implemented for a conformal coating used to prevent tin whisker growth. The advanced blister testing with predefined area test method was then extended by employing a numerical method for evaluating the adhesion strength when the polymer’s film properties are unknown.

Slot Film Cooling: A Comprehensive Experimental Characterization

When components of a propulsion system are exposed to elevated flow temperatures there is a risk for catastrophic failure if the components are not properly protected from the thermal loads. Among several strategies, slot film cooling is one of the most commonly used, yet poorly understood active cooling techniques. Tangential injection of a relatively cool fluid layer protects the surface(s) in question, but the turbulent mixing between the hot mainstream and cooler film along with the presence of the wall presents an inherently complex problem where kinematics, thermal transport and multimodal heat transfer are coupled. Furthermore, new propulsion designs rely heavily on CFD analysis to verify their viability. These CFD models require validation of their results, and the current literature does not provide a comprehensive data set for film cooling that meets all the demands for proper validation, namely a comprehensive (kinematic, thermal and boundary condition data) data set obtained over a wide range of conditions. This body of work aims at solving the fundamental issue of validation by providing high quality comprehensive film cooling data (kinematics, thermal mixing, heat transfer). 3 distinct velocity ratios (VR=uc/u∞) are examined corresponding to wall-wake (VR~0.5), min-shear (VR ~ 1.0), and wall-jet (VR~2.0) type flows at injection, while the temperature ratio TR= T∞/Tc is approximately 1.5 for all cases. Turbulence intensities at injection are 2-4% for the mainstream (urms/u∞, vrms/u∞,), and on the order of 8-10% for the coolant (urms/uc, vrms/uc,). A special emphasis is placed on inlet characterization, since inlet data in the literature is often incomplete or is of relatively low quality for CFD development. The data reveals that min-shear injection provides the best performance, followed by the wall-jet. The wall-wake case is comparably poor in performance. The comprehensive data suggests that this relative performance is due to the mixing strength of each case, as well as the location of regions of strong mixing with respect to the wall. Kinematic and thermal data show that strong mixing occurs in the wall-jet away from the wall (y/s>1), while strong mixing in the wall-wake occurs much closer to the wall (y/s<1). Min-shear cases exhibit noticeably weaker mixing confined to about y/s=1. Additionally to these general observations, the experimental data obtained in this work is analyzed to reveal scaling laws for the inlets, near-wall scaling, detecting and characterizing coherent structures in the flow as well as to provide data reduction strategies for comparison to CFD models (RANS and LES).

Interaction of Acoustic Waves with a Laminar Line-Flame

A systematic study was conducted to elucidate the effects of acoustic perturbations on laminar diffusion line-flames and the conditions required to cause acoustically-driven extinction. Flames were produced from the fuels n-pentane, n-hexane, n-heptane, n-octane, and JP-8, using fuel-laden wicks. The wicks were housed inside of a burner whose geometry produced flames that approximated a two dimensional flame sheet. The acoustics utilized ranged in frequency between 30-50 Hz and acoustic pressures between 5-50 Pa. The unperturbed mass loss rate and flame height of the alkanes were studied, and they were found to scale in a linear manner consistent with Burke-Schumann. The mass loss rate of hexane-fueled flames experiencing acoustic perturbations was then studied. It was found that the strongest influence on the mass loss rate was the magnitude of oscillatory air movement experienced by the flame. Finally, acoustic perturbations were imposed on flames using all fuels to determine acoustic extinction criterion. Using the data collected, a model was developed which characterized the acoustic conditions required to cause flame extinction. The model was based on the ratio of an acoustic Nusselt Number to the Spalding B Number of the fuel, and it was found that at the minimum speaker power required to cause extinction this ratio was a constant. Furthermore, it was found that at conditions where the ratio was below this constant, a flame could still exist; at conditions where the ratio was greater than or equal to this constant, flame extinction always occurred.

STRAINING ENGINEERING OF GRAPHENE AND GRAPHENE-BASED NANOSTRUCTURES

Graphene has emerged as an extraordinary material with its capability to accommodate an array of remarkable electronic, mechanical and chemical properties. Extra-large surface-to-volume ratio renders graphene a highly flexible morphology, giving rise to intriguing observations such as ripples, wrinkles and folds as well as the potential to transform into other novel carbon nanostructures. Ultra-thin, mechanically tough, electrically conductive graphene films promise to enable a wealth of possible applications ranging from hydrogen storage scaffolds, electronic transistors, to bottom-up material designs. Enthusiasm for graphene-based applications aside, there are still significant challenges to their realization, largely due to the difficulty of precisely controlling the graphene properties. Controlling the graphene morphology over large areas is crucial in enabling future graphene-based applications and material design. This dissertation aims to shed lights on potential mechanisms to actively manipulate the graphene morphology and properties and therefore enable the material design principle that delivers desirable mechanical and electronic functionalities of graphene and its derivatives.

Topics in equilibrium and nonequilibrium thermodynamics: computing crystalline free energies and engineering Maxwell’s demon.

This dissertation covers two separate topics in statistical physics. The first part of the dissertation focuses on computational methods of obtaining the free energies (or partition functions) of crystalline solids. We describe a method to compute the Helmholtz free energy of a crystalline solid by direct evaluation of the partition function. In the many-dimensional conformation space of all possible arrangements of N particles inside a periodic box, the energy landscape consists of localized islands corresponding to different solid phases. Calculating the partition function for a specific phase involves integrating over the corresponding island. Introducing a natural order parameter that quantifies the net displacement of particles from lattices sites, we write the partition function in terms of a one-dimensional integral along the order parameter, and evaluate this integral using umbrella sampling. We validate the method by computing free energies of both face-centered cubic (FCC) and hexagonal close-packed (HCP) hard sphere crystals with a precision of $10^{-5}k_BT$ per particle. In developing the numerical method, we find several scaling properties of crystalline solids in the thermodynamic limit. Using these scaling properties, we derive an explicit asymptotic formula for the free energy per particle in the thermodynamic limit. In addition, we describe several changes of coordinates that can be used to separate internal degrees of freedom from external, translational degrees of freedom. The second part of the dissertation focuses on engineering idealized physical devices that work as Maxwell's demon. We describe two autonomous mechanical devices that extract energy from a single heat bath and convert it into work, while writing information onto memory registers. Additionally, both devices can operate as Landauer's eraser, namely they can erase information from a memory register, while energy is dissipated into the heat bath. The phase diagrams and the efficiencies of the two models are solved and analyzed. These two models provide concrete physical illustrations of the thermodynamic consequences of information processing.

THE ROLE OF THE MECHANICAL ENVIRONMENT ON CANCER CELL TRANSMIGRATION AND MRNA LOCALIZATION

Most cancer-related deaths are due to metastasis formation, the ability of cancer cells to break away from the primary tumor site, transmigrate through the endothelium, and form secondary tumors in distant areas. Many studies have identified links between the mechanical properties of the cellular microenvironment and the behavior of cancer cells. Cells may experience heterogeneous microenvironments of varying stiffness during tumor progression, transmigration, and invasion into the basement membrane. In addition to mechanical factors, the localization of RNAs to lamellipodial regions has been proposed to play an important part in metastasis. This dissertation provides a quantitative evaluation of the biophysical effects on cancer cell transmigration and RNA localization. In the first part of this dissertation, we sought to compare cancer cell and leukocyte transmigration and investigate the impact of matrix stiffness on transmigration process. We found that cancer cell transmigration includes an additional step, ‘incorporation’, into the endothelial cell (EC) monolayer. During this phase, cancer cells physically displace ECs and spread into the monolayer. Furthermore, the effects of subendothelial matrix stiffness and endothelial activation on cancer cell incorporation are cell-specific, a notable difference from the process by which leukocytes transmigrate. Collectively, our results provide mechanistic insights into tumor cell extravasation and demonstrate that incorporation into the endothelium is one of the earliest steps. In the next part of this work, we investigated how matrix stiffness impacts RNA localization and its relevance to cancer metastasis. In migrating cells, the tumor suppressor protein, adenomatous polyposis coli (APC) targets RNAs to cellular protrusions. We observed that increasing stiffness promotes the peripheral localization of these APC-dependent RNAs and that cellular contractility plays a role in regulating this pathway. We next investigated the mechanism underlying the effect of substrate stiffness and cellular contractility. We found that contractility drives localization of RNAs to protrusions through modulation of detyrosinated microtubules, a network of modified microtubules that associate with, and are required for localization of APC-dependent RNAs. These results raise the possibility that as the matrix environment becomes stiffer during tumor progression, it promotes the localization of RNAs and ultimately induces a metastatic phenotype.