ABSTRACT. The combination of biochemical and mechanical stimulus regulates cellular activity and ultimately, the correct functioning of human beings. Whilst mechanical views for studying biological processes were casting aside, recent attention have been drawn toward this point. In this direction, a widespread practice for the study of cellular behavior against mechanical cues relies on the use of different substrates. Traditionally, this has been carried out by inducing static changes in substrate’s culture mechanical properties. In this study we present a new device able to dynamically change the mechanical characteristic of a substrate by means of an electroactive piezoelectric polymer. The biocompatible arrangement is calibrated by an interferometric system; and used to study cellular responses entailing proliferation, migration and morphological changes. Our study suggest that vibrations at low frequencies enhance migrations rates and proliferation cue in an epithelial cell line, whereas a frequency of 80 Hz produces contrary effects. Morphology is considerably affected under both dynamic conditions

ABSTRACT. Osteoarthritis (OA) is a debilitating joint disease caused by chronic inflammation and wear and tear of articular cartilage. It is the most common form of arthritis, affecting 27+ million people in the United States. Sustained delivery of platelet-rich plasma (PRP) is a promising OA therapy, with the potential to slow, reverse or halt disease progression. However, when PRP is delivered via intra-articular injection, it is cleared from the joint rapidly, diminishing treatment efficacy. We hypothesize that a delivery vehicle for sustained PRP release will improve outcomes. We fabricated biodegradable polyethylene glycol (PEG) microspheres of varying sizes via microfluidics by manipulating several key parameters. The PRP-PEG microspheres were characterized for size and size distribution, PRP release and degradation in synovial fluid. In-vitro studies on treatment efficacy were performed and in vivo studies are ongoing. PRP-PEG microspheres had narrow size distribution under all tested flow rate combinations. The microsphere diameter decreased with increase in oil flow rate and decrease in PEG flow rate. Microsphere diameter increased 1.36-times upon washing. PRP was released over ~11 days with 40% release. PRP-PEG releasate from hydrogel slabs had beneficial effect on chondrocyte proliferation and certain gene expression profiles. Microspheres were stable in synovial fluid for ~72 h, but degradation in synovial fluid was significantly faster than in PBS. Degradation in the presence of PRP in the PEG microspheres was significantly slower than in the absence of PRP in vitro and in vivo. Treatment efficacy and toxicity testing in a mouse model of knee OA is ongoing.

ABSTRACT. Decellularized extracellular matrix can aid to recreate the host environment due to the similar protein structure to native muscle. The sponge-like structure of cryogels is macroporous and ideal for its rapid swelling along with the ability to withstand repeated cyclic loads without major deformation or loss of structural integrity. This study incorporated cryogels with decellularized muscle in varying concentrations and were compared to gelatin cryogels. The glutaraldehyde crosslinker was varied for improved cryogel structure. Ultimate and cyclic testing shows decellularized ECM makes a stronger scaffold with a notable higher peak stress and modulus of elasticity compared to gelatin counterparts. Decellularized ECM cryogels are more resilient to stress as shown by smaller hysteresis values. The swell ratio also decreases with use of decellularized ECM showing that the cryogel is less porous than the gelatin cryogels. These factors are most likely due to a higher protein content which creates a more structurally intact scaffold. When the glutaraldehyde content was increased, the protein content, mechanical properties, and swell ratio also increased. Elution from the decellularized ECM increased cell proliferation compared to their gelatin counterparts after 7 days. It is expected that the decellularized ECM scaffold will thus promote cellular infiltration and activity. The combination of these properties shows its potential as a porous scaffold strong enough to withstand cyclic loading from muscle while simultaneously promoting cellular activity.

ABSTRACT. Many tumors express thymidine phosphorylase (TP) with various levels, however due to tumor heterogeneity the amount of TP is usually not enough to convert prodrug 5’-DFUR to toxic drug 5-FU. In this study, TP plasmids were first delivered to cancer cells and treated with prodrug 5’-DFUR. TP level and prodrug conversion were confirmed by enzyme activity tests. The TP-transfected cancer cells treated with 5’-DFUR revealed higher death rate than the non-transfected ones. Optimal conditions of nonviral gene delivery of TP-GFP were tested by transfecting A549 cells and human mesenchymal stem cells (MSC) under different inoculation cell numbers, medium conditions (serum/serum-free), DNA amounts, and N/P ratios. Because MSC is tumor-tropic and low immunogenic, it is good carrier to deliver drugs. TP plasmid was delivery to MSCs and co-culture with A549 cells. Two ratios of MSCs to A549 cells in the co-cultured wells were tested. Our results showed that 2:1 ratio led to A549 cell death on day 2, in comparison with cell death on day 3 for 1:1 ratio. Moreover, the eradication of A549 cancer cells was shown after 5 days for 2:1 ratio of MSCs to A549 cells for prodrug therapy.

ABSTRACT. Introduction: Microspheres have been used in many embolization applications including intravascular treatment for Liver Cancer and Benign Prostatic Hyperplasia. These microspheres are generally not visible and are not degradable which prevents repeat treatment or degradation tracking. The goal of the study was to create degradable microspheres that are radiopaque.

Materials and Methods: A microfluidic setup was used to create the microspheres. A 20% PEG precursor solution was made with PEG Acrylate and PEG Dithiol in a 1:1 molar ratio between acrylate and thiol groups. To this solution Barium Sulfate was added to achieve a 10% w/v. Two syringes perpendicular to each other using a tee junction, one with Barium-loaded PEG solution and one with Kirkland Olive oil, were placed on two separate micro pumps set at 10μl/min for the PEG phase and 1000μl/min for the oil phase. Microspheres were created due to the immiscible nature of the two liquids. The spheres were then collected in an oil bath and imaged.

Results and Discussion: A portion microspheres were dried in microfuge tubes and remainder were injected subcutaneously into a live mouse. After rehydration of dry microspheres, MicroCT analysis showed that Barium Sulfate was encapsulated evenly in several microspheres but not in others. Even distribution is possible but the manufacturing process needs refinement. Cone beam CT analysis of the mouse, after sacrifice, showed clearly visible barium-loaded microspheres. This formulation can therefore be successfully imaged in a clinical setting.

ABSTRACT. Air-gap electrospinning is a method for consistently producing highly aligned polymer nanofibers by electrospinning across two positively charged plates. In this study, a modified air-gap electrospinning setup was exploited to wrap highly aligned polycaprolactone polymer nanofibers around individual 1393 bioactive glass microfibers continuously. This resulted in a synthetic structure similar to osteons, which is the repeating unit of cortical bone. By varying the disc diameter, charge, rotation speed, and the location on the glass fiber, polymer fibers that were wrapped at angles between 10-30 to the glass fiber were chosen, although fibers wrapped as large as 45-90 were possible. Compared to randomly aligned electrospun fibers, this scaffold is expected to increase migration along the fiber direction, as shown in recent work regarding nerve and muscle regeneration along aligned fibers. There was no change in the fiber diameter, although the porosity decreased from 90% to 80% due to consolidation of the aligned fibers during wrapping. Encapsulating the glass with polymer nanofibers caused viscoelastic deformation during 3-point bending, but no statistical change in peak stress. Scaffold degradation was not cytotoxic. The aligned polymer fibers demonstrated unidirectional cell alignment, indicating increased cell movement speed. It is predicted that the bioactive glass microfibers will convert into hollow fibers, allowing blood vessel ingrowths, while the aligned polymer fibers will stimulate cell migration into the scaffold. This design is expected to have staggered release of synthetic and organic dopants, with controllable degradation rates based off the compositions used for the glass and polymer.

ABSTRACT. Fibrillar collagen, a primary component of musculoskeletal tissues, is formed by a complex, multistep process in the extracellular space. The rate-limiting step for collagen fibril formation is the removal of the C-terminus domain from the secreted procollagen molecule by the actions of bone morphogenetic protein-1 (BMP-1). Because procollagen conversion and fibril assembly occur slowly during in vitro culture, there is substantial loss of secreted procollagen into the medium and limited collagen deposition around cells, representing a major challenge for ex vivo musculoskeletal tissue engineering approaches. Macromolecular crowding (MMC) approaches can enhance collagen deposition in 2D cultures of various cell types, with effectiveness of the MMC reagent depending on macromolecular charge, size, and concentration. For example, we have shown that use of negatively-charged dextran sulfate in cultures of MG-63 osteoblast-like cells increased collagen deposition by 80% compared to control cultures, while collagen deposition with use of neutral dextran was unchanged. How the MMCs enhance procollagen conversion and collagen accumulation is poorly understood. Here, we use fluorescence correlation spectroscopy, which allows for non-invasive, real-time measurement of diffusivity and interactions, to evaluate how MMC charge and concentration affects the diffusivity of a model protein. We use dextran sulfate and neutral dextran as the crowders and bovine serum albumin as the model protein. Next, we will evaluate diffusion of type II procollagen and BMP-1 in the presence of dextran sulfate and neutral dextran of varying concentration and assess whether transport-related effects contribute to the enhancement of procollagen processing observed in the presence of MMCs.

ABSTRACT. Multiple myeloma (MM) is a plasma B-cell hematologic malignancy that causes significant skeletal morbidity. CD38 is a type II transmembrane glycoprotein over-expressed in myeloma cells. Daratumumab is an FDA approved high-affinity monoclonal antibody targeting CD38 that has shown promising therapeutic efficacy in double refractory MM patients. However, heterogeneity in therapy response remains a major challenge, thus necessitating stratification strategies to minimize off-target toxicities and improve quality of life. Here, we have developed and validated a daratumumab-conjugated near-infrared (NIR) fluorescent contrast agent for in vivo and ex vivo spatial localization of CD38 positive MM tumors. NIR imaging provides improved depth penetration due to reduced absorbance by endogenous chromophores, allowing economical and high-throughput preclinical imaging of MM tumors. We hypothesize that the enhanced specific expression of CD38 glycoprotein on malignant plasma cells will favor increased IRDye800-daratumumab uptake and allow for clinically impactful fluorescence imaging for therapeutic planning as an ex vivo companion diagnostic. These studies will also contribute toward technological advancements in diagnostic and therapeutic monoclonal antibody-based pharmaceutical development for MM and lymphoid tumors in general.

ABSTRACT. Acute Lymphoblastic leukemia is a malignant disease characterized by the accumulation of lymphocytes in the bone marrow. The white blood cells (WBCs) analysis allows the detection of acute Lymphoblastic leukemia. In this work, we introduce an automated method for blood cells classification and analysis in microscopic images. In this work, we apply different segmentation techniques to isolate WBCs from microscopic images. Then, several features such as shape, color, and texture are extracted. In this work, we introduce the flow of the steps that can be applied to automate the detection techniques. This automated analysis gained a high accuracy that is used to detect Acute Lymphoblastic leukemia. The extracted features are used to train different classification models in order to determine the segmentation techniques accuracy and the performance of using several classification models. The automated approach represents a medical tool used to avoid the numerous drawbacks associated with manual observation. These automated image techniques could open the door for more finding in the biomedical researches.

ABSTRACT. Obesity has a high relation with plenty of diseases and becomes one of the biggest concerns in current society. Excessive energy storage as fat in adipose tissue leads to the tremendous increasing prevalence of obesity-related diseases. In this study, the effects of two important receptors, (G-protein coupled receptor 120 (GPR120) and cluster of differentiation 36 (CD36), on fatty acid sensing and uptake process were analyzed by Surface Enhanced Raman Spectroscopy (SERS) at single living cell level. Two antibody specific nanoprobes (NPs) as molecular sensors were synthesized and applied to specifically detect the expression levels of GPR120 and CD36 on cell membrane by monitoring of unique SERS peaks upon the addition of fatty acid linoleic acid (LA). Real-time monitoring of interaction between the SRES probes and the receptors provided extracellular recognition process upon fatty acid stimuli, which supplemented intracellular Ca2+ activity measurement by calcium imaging system. Receptor internalization process was occurred in both immortalized and fresh taste bud cell, while not seen in HEK293 cell. These findings suggest GPR120 acts as a primary receptor to fatty acid and works synergistically with CD36 to transfer signals inside cell, further activating the process of fatty acid uptake. Moreover, we expanded the application of NPs into mouse tongue tissue sample, which successfully proved the feasibility of our developed sensing probe as a promising tool in animal model. It might provide more insights for human in dietary selectivity and obesity management.

ABSTRACT. For many optical and electronic devices employing thin films as a surface coating, the coating’s chemical, mechanical, and structural stability when exposed to complex environments are key factors that determine device lifetime and impact the device’s performance metrics. For instance, repeated or prolonged exposure of polymeric coatings, such as poly methyl (methacrylate) (PMMA), to water and other chemicals can lead to the potential dissolution of the coating or to the accumulation of materials and spot defects, as well as to increased surface roughness; this may result in changes to the coating’s refractive index and thickness and may reduce the coated component’s lifetime. However, determining the ability of thin films or coatings to withstand harsh working conditions is rarely studied, primarily because the standard characterization techniques, such as ellipsometry, are limited in terms of the types of materials they can address. OTPAS overcomes this fundamental limitation by using a pulsed evanescent field to induce a photoacoustic effect and spectroscopically probe nanomaterials’ characteristics, providing early signs of degradation. Here, we demonstrate OTPAS’s ability to track the dissolution of microscale PMMA coatings in an etching solution that dissolves small amounts of PMMA over time and to monitor the changes in coating thickness and refractive index; the results are compared with standard ellipsometric protocols to showcase the capabilities of the technique. This work represents the first example of OTPAS’s use in evaluating materials degradation.

ABSTRACT. Globally, there are approximately eleven million injuries related to burns that require care each year. Due to the current methods used for diagnosis, only about 66% of wounds are correctly diagnosed. Misdiagnosis can lead to unnecessary treatments, costing patients and burn wound specialists time and money. Alternative methods of wound analysis have proven to encompass a variety of limitations, presenting the need for a method that provides quantitative information. We have developed a waveguide-mediated hand-held device that uses backward-mode optoacoustic computed tomography (OACT) to image synthetic burn wound tissue, which prevents signal destruction while retaining sensitivity. We are implementing a graphical processing unit to accelerate imaging speed. To increase the energy density provided to the tissue we designed an optical train using Zemax OpticStudio. We have successfully simulated a spot size suitable for our waveguide entrance. We will be building it in lab to allow for further testing using an optical fiber with a larger core diameter. Our goal is to reach an energy density of 18 mJ/cm2 to image a burn wound on human tissue. We will be altering the settings of the digital filter to increase the signal to noise ratio, reducing the number of averages taken of the signals. This device presents the opportunity for imaging wounds of human tissue to ultimately provide a method for clinicians to more accurately diagnose burns.

ABSTRACT. World population is expected to reach nearly 10 billion by 2050. In 30 years we would add more than total number of people currently living in India and China combined. We are making farming more precise and smarter. But will that be enough, especially with the growing scarcity of water and land for a world in which 70% people ive in urban areas away from from farms.

Since the time human first dropped seeds on earth surface to produce more, we have been doing the same thing - till, seed/plant, tend and harvest - called farming. We innovated new ways but they all focus on improving our ability to get greater harvest from a unit area of land.

The grand question today is: Will we be able to increase farm productivity with diminishing resources by to produce 50% more food in 30 years to match 2050 world food demand?

Other than hunting, food come from living systems produced by conventional farming. Farming is the regeneration of biological materials for food. Advances in biology and information sciences make one wonder whether new ways could be created for regenerating (synthesizing) biological materials for food. Engineers creates things that have never existed before, so why not explore new possibilities? Is this the time to create something new, designed to predetermined specifications and taking fewer resources?

This presentation will discuss these challenges with the hope of influencing others to join this conversation and explore new biology inspired ways for "producing" food.

ABSTRACT. The present study evaluated the organosolv pulp of wheat straw, CIRNO variety, cultivated in the Yaqui Valley during the 2016-2017 production cycle, to determine if it is a good substrate for the production of fermentable sugars. 36 native edaphic fungi from the Yaqui Valley and the reference fungus Trichoderma reesei ATCC 56765 (control) were evaluated to select the fungal strain that presented the best enzymatic cellulolytic activity and finally used it in enzymatic hydrolysis. Through a submerged bioprocess, the TRQ90 native fungus presented a greater potential compared to the other native fungal strains, obtaining enzymatic values of 11.4 ± 2 FPU/mg. Furthermore, the straw cellulose pulp proved to be a viable raw material for the production of fermentable sugars since it had a value exceeding the filter paper sugars production by 0.75 ± 0.2 mg/ml. Strain TRQ90 presented superior values before the control strain. It is recommended to optimize the processes of enzyme production and extraction, together with the enzymatic hydrolysis process, which will improve the production of fermentable sugars to contribute with the development of bioenergies through the use of native fungi and wheat straw cellulose pulp from Yaqui Valley.

ABSTRACT. Globally, approximately one-third of all food is wasted. The economic and environmental consequences of food waste are dire: in 2017, the National Resources Defense Council estimated that every year, food waste costs the United States 218 billion dollars, accounts for 21% of landfill material, and generates nearly 3% of all US greenhouse gas emissions. Mostly, food waste occurs during the production stages of food preparation, making it a promising starting point for researchers. A major challenge, colonies of bacteria that grow on food-processing surfaces cause food spoilage. To address this challenge, previous workers have successfully developed photocatalytic antimicrobial thin-films with various metal-oxides, albeit largely without consideration to industrial needs, like ease-of-processing and durability. We further the effort by growing chemically-inert films of TiO2 with enhanced mechanical strength onto type 316L stainless steel, a common food processing surface material. Our results include measurements of film quality, reactivity, surface adhesion strength, and mechanical strength to which we compare results from previous works. Lastly, the films are tested against common fouling bacteria, like listeria, salmonella, and E. coli with promising results.

ABSTRACT. Cancer is the second leading cause of death worldwide. Melanoma and nonmelanoma skin cancer (NMSC), one of the most common type of cancer, have been accounted for 1.04 million cases in 2018. Exposure of skin to intensive ultraviolet radiation has been associated with melanoma and NMSC. The UV component of sunlight is a major physical carcinogen causing cancer in humans. Reducing exposure to ultraviolet radiation can significantly prevent new incidences of NMSC. Therefore, quantifying the amount of UV exposure in one’s daily life helps monitoring the health situation for the sake of skin cancer. In this research, an on-chip all-silk fibroin whispering gallery mode microresonator fabricated by soft lithography method was developed for ultraviolet sensing. The fabrication and testing of this device have been experimentally demonstrated. The quality factors of the silk microresonators are on the order of 105, which provides a high sensitivity for UV detection. Thanks to the nature of the biocompatibility and biodegradability of structural protein (i.e. silk fibroin), these optical sensors demonstrate possibilities for biomedical and biophotonics applications in the future.

ABSTRACT. Listeria monocytogenes is a deadly foodborne pathogen that has led to multiple outbreaks and is responsible for a high mortality rate. The conventional methods used for Listeria detection include standard microbiology or biochemical procedures, which require time-consuming sample pretreatment techniques that can take up to 72 hours. There is a growing need for methodologies that reduce the test time for Listeria detection from days to an hour or less, while also allowing for scalability and high-volume processing. The challenge lies in pretreating large volumes of food samples to generate a sufficient analyte for detection tests within a short time period, without compromising the selectivity of the test. We have previously worked on the development of separation and detection methods for bacterial cells [1, 2]. This work describes a magnetophoretic microfluidic device that can selectively isolate and concentrate tagged Listeria monocytogenes from food matrices. Listeria cells will be tagged using bacteriophage-functionalized magnetic particles. A multi-chamber microfluidic device will be used in combination with an external magnet to trap and therefore separate magnetically-tagged Listeria cells from food matrices. This method of concentration has applications in rapid diagnostic systems such as biosensors for the detection of pathogenic bacteria in food samples.

1. Ghuman, A., et al., Magnetic Separation of Food-Borne Pathogens in a Microfluidic Device, in IBE 2018 Annual Conference. 2018. 2. Zhou, Y., et al., Isolation and Separation of Listeria monocytogenes Using Bacteriophage P100-Modified Magnetic Particles. Colloids and Surfaces B: Biointerfaces, 2018. Under Review.

ABSTRACT. Bacterial foodborne illness affects approximately 9.4 million people annually in the United States. One prevalent pathogen is Listeria monocytogenes, which can propagate in temperatures from -1.5 to 50°C and pH 4.3 to 9.6. Listeria has been associated with recalls in fresh produce to frozen food. The overall goal of this research is to develop low cost, laser carbonized biosensors for rapid, point of use detection of L. monocytogenes in large sample volumes (225 mL according to FDA testing criteria) via vacuum driven flow directly through electrodes. Flexible laser scribed graphene (LSG) electrodes were engineered by carbonization of a polyimide film using a low-cost UV laser. Laser pulse width was optimized by measurements of electroactive surface area, heterogenous electron transfer kinetics, and charge transfer resistance. Based on electrochemical performance, the optimal laser pulse width was found to be 40 ms. LSG electrodes were metallized and biofunctionalized with a L. monocytogenes specific DNA aptamer. The current limit of detection (LOD) for L. monocytogenes in 10 mL sample volumes is 10 CFU/mL in 20 minutes using electrochemical impedance spectroscopy. Vacuum driven flow of large analytical samples directly through porous nanohybrid electrodes is implemented to achieve low limits of detection within 4-6 hours of total processing time including sample preparation, incubation, testing, and analysis. Traditional detection methods typically take 48-72 hours. The developed LSG biosensors are easily reproducible with a high degree of customizability, demonstrating a new analytical platform for rapid food safety monitoring and potential use in hospitals, disease tracking, and ecosystem health.

ABSTRACT. Methicillin-resistant Staphylococcus aureus (MRSA) is a bacterial strain that is resistant to many antibiotics. In hospitals and other healthcare environments, MRSA, if not treated quickly, can cause severe complications such as bloodstream infections, pneumonia, and surgical site infections leading to sepsis and death. Current detection methods for MRSA either require trained personal to operate and perform expensive tests or are too time-consuming. Electrochemical biosensors offer unique advantages such as high sensitivity and rapid results. In this study, we developed an electrochemical biosensor for selective detection of MRSA strain USA300 with high sensitivity. The biosensor could potentially be integrated into lab-on-a-chip platforms for point of care use.

ABSTRACT. Preterm birth is the leading cause of neonatal death, however no accurate methods for predicting preterm birth exist. Detection of early changes in the cervix, an organ that must biochemically remodel to allow for passage of a fetus, could be a useful predictor for preterm birth risk. Researchers have employed light-based methods to monitor biochemical changes in the cervix during pregnancy, however, all optical approaches developed thus far require patients to undergo a speculum exam, which many patients find uncomfortable and is not standard practice during prenatal care. Development of a visually guided optical tool is presented that measures the cervix via bimanual exam, a routine procedure performed by Women’s health professionals during prenatal visits and labor for monitoring of the cervix. The tool utilizes a Raman spectroscopy probe for biochemical monitoring of the cervix and a camera for visualizing measurement location to ensure it is clean of biological fluids. The newly developed probe was tested first in lower reproductive tract mannequins followed by fifteen patients receiving Obstetric and Gynecological care. Results acquired with and without a speculum revealed similar spectra, demonstrating that Raman spectral information is conserved when using the visually guided probe. Additionally, the majority of patients stated that the new device reduced discomfort. In conclusion, the speculum-free visually-guided probe was successfully integrated with standard prenatal procedures and obtained high quality Raman measurement of the cervix. This work overcomes a large barrier in clinical translation of light-based t methods for quantitative cervical assessment during pregnancy.

ABSTRACT. During pregnancy, the cervix must undergo significant biochemical remodeling to allow for successful labor and delivery. In vivo Raman spectroscopy is an optical technique that can be used to investigate the biochemical composition of tissue longitudinally and non-invasively in humans and has been utilized to measure physiology and disease states in a variety of medical applications. We hypothesized that Raman spectral signatures reflect known biochemical remodeling. To test this hypothesis, over 60 patients receiving prenatal care were recruited and Raman spectra were measured using an in vivo Raman system with an optic fiber probe throughout pregnancy. The data was analyzed multiple ways, including the development of a longitudinal spectral model and fitting a non-negative least squares model based on pure Raman spectra from actin, blood, collagen type I, cholesterol, glycogen, and water. Least squares analysis revealed a decrease in collagen signal (p<0.05) throughout pregnancy, which agrees with extensive reports on extracellular matrix breakdown. In addition, signatures corresponding to blood significantly increased, in agreement with reports revealing increased vascularity as the cervix prepares for delivery. This study has demonstrated sensitivity to known biochemical dynamics that occur during cervical remodeling non-invasively, and identified patient variables including body mass index and obstetric history that significantly effect Raman signals. Raman spectroscopy has the potential to aid global efforts to close the gaps in our understanding of cervical maturation, and ultimately reduce the incidence, morbidity, and mortality caused by preterm birth.

ABSTRACT. In many real-world situations, one seeks to detect viable microorganisms present in samples where dead microorganisms are known/expected to be present. Examples include (1) detection of viable Mycobacterium tuberculosis (organism causing Tuberculosis) in sputum of patients suspected of having active infection due to dormant M.tuberculosis presence or dead M.tuberculosis left behind after previous treatment, (2) bacteria (ex. Lactic-acid-bacteria) that survived decontamination efforts and present in inoculums of yeast cultures used for ethanol production. Because DNA-based methods like ELISA give false-positives, indirect metabolism-based approaches are typically employed. Here, one detects bacterial metabolism/growth signature (either generic changes such as pH, conductivity changes or specific metabolite production). They are limited by metabolism rate and doubling-time of microorganisms, and thus have long times-to-detection(TTD). In contrast, we ask, “can they be killed”? Since only living entities can be killed, an answer in the affirmative confirms presence of live entities/microorganisms. Similar to detection based on metabolism, we can choose either a generic or species-specific killing-agent to serve as signatures for presence of target species. Real-time detection of bacterial cell death is achieved using Microchannel-Electrical Impedance Spectroscopy(m-EIS). Briefly, relies on the fact that live cells have non-zero membrane-potential, and hence when exposed to high-frequency AC-field, accumulate charges at membranes, contributing to suspension’s bulk-capacitance. This charge storage does not occur at dead cell membranes across which no potential-difference exists. Doing so makes our TTD dependent on how fast they are killed (usually within minutes) and/or detection threshold(~1000CFU/ml), making our TTDs dramatically shorter (hours instead of weeks for TB).

ABSTRACT. The spread of multi-drug resistant tuberculosis (MDR-TB) is a global public-health challenge. Because not all biological-mechanisms of resistance are known, culture-based (phenotypic) drug susceptibility testing provides important information that influences clinical decision-making. Current phenotypic tests typically require pre-culture step taking 2-4weeks, followed by 10-14days to confirm growth or lack thereof. We present a 2-step method to obtain DST results within 3days of sample collection. First-step involves selectively concentrating live mycobacterial-cells present in relatively large volumes of sputum using commercially-available magnetic-nanoparticles (MNPs) into smaller volumes, thereby bypassing the need for pre-culture. Second-step involves using microchannel-Electrical Impedance Spectroscopy (m-EIS) to monitor multiple aliquots of small volumes (~10μL) of suspension containing mycobacteria, MNPs and candidate drugs to determine cell growth/death/stasis under the conditions tested. m-EIS determines solution “bulk-capacitance”, a parameter proportional to the number of live bacteria in suspension. Thus, we can detect cell death (bactericidal action of the drug) in addition to cell growth. We demonstrate proof-of-principle using M.bovis BCG and M.smegmatis suspended in artificial sputum. Loads of ~2000–10,000CFU of mycobacteria were extracted from ~5ml of artificial sputum during decontamination process with 84%-100% efficiencies. Subsequently, suspensions containing ~105CFU/ml of mycobacteria with 10mg/ml of MNPs were monitored in the presence of bacteriostatic/bactericidal drugs at concentrations below/at/above known Minimum Inhibitory Concentration (MIC) values. m-EIS data showed data consistent with growth/death/stasis as expected and/or recorded using plate-counts. Electrical signals of death were visible as early as 3hours, and growth in < 3days for all samples, thus allowing us to perform DST in < 3days.

ABSTRACT. One metric to monitor plant growth, especially in greenhouses, is the area coverage by the plant. There is a need to automate this process, and current approaches are widely based on visible light imaging. The challenge is a reliable separation of the leaf area from the background, because the latter can include a wide variety of materials (e.g., soil, metal, concrete, or even dead plant matter). The use of chlorophyll fluorescence advertises itself, because the plant shows a high contrast with fluorescent leaves over a dark background. The major cause of false-positive regions in this case are algae. The two most crucial steps for the removal of algae fluorescence is to use a white-light reference for intensity normalization, and to use texture classification to separate the generally irregular algae texture from the more homogeneous leaf fluorescence. We present examples of calibrated area measurements, but also highlight challenges caused by partly shaded leaves and by diseases, such as tip burn.

ABSTRACT. Methyl salicylate (MeSa) is an important plant volatile organic compound (VOC) released by plants under biotic stress events. Therefore, detection MeSa could help realize early detection of plant diseases before symptoms appear. It also has a profound significance for precision agriculture for maintaining effective use of pesticides and fungicides1. In this work, the design and development of an electrochemical biosensor for the detection of MeSa released by plants was conducted 2, 3. A 3D printed VOC sample collection device is designed, fabricated and used for VOC collection and pre-concentration 4. In addition, the detection of MeSa after collection and pre-concentration is performed by a lab based microfluidic device.

ABSTRACT. The fluorescence emitted by chlorophyll in plant leaves provides information about how efficient it uses the incident light, but also about its health. A primary goal of this project is to determine optimum light levels for supplemental lighting. We have shown previously that an automated biofeedback system on the basis of chlorophyll fluorescence translates into significant costs savings. Instruments that measure chlorophyll fluorescence exist, but are too expensive to enable their widespread adoption. Therefore, there is a need to develop a low-cost instrument, and our goal was to keep materials costs below $200. We presented such a device before, but also acknowledged some challenges, such as insufficient light intensity and nonlinear sensor behavior. Since then, we have improved the design and remedied the shortcomings. The second iteration of the chlorophyll fluorometer uses laser excitation and an avalanche photodiode (APD) for improved sensitivity. With the laser, excitation-path optics can be minimized for cost reduction. A special integration circuit for the APD corrects nonlinear behavior and led to a 10-fold improvement of the signal-to-noise ratio over the first generation. Finally, we also integrated actinic light control into the device to use it as a standalone biofeedback system.

ABSTRACT. Soybean is a valuable crop as one of the major protein sources for human diet, feed for livestock and aquaculture, and as an oil seed crop. Soybean breeding is an on-going program to produce high-yielding soybean varieties that could sustain the future needs. Phenotypic data from soybean such as plant height, lodging, and maturity date are correlated with the soybean genetic traits and used for breeding selection. These data are collected by field measurement and visual inspection from experienced breeders. Remote sensing techniques especially using unmanned aerial system (UAS) has been adopted lately in plant phenotyping in field condition using plant reflectance, vegetation indices (VIs), canopy temperature, and estimated plant height. The main objective of this study is to evaluate the potential use of UAS-based imaging system in making soybean breeding line selection based on plant reflectance, VIs, temperature, and estimated soybean yield. Various imaging sensors (a high-resolution digital camera for color images, a multispectral camera and a thermal camera) were mounted on an unmanned aerial vehicle (UAV) to take aerial images on 288 soybean breeding plots during their growth stages of R1 and R3. VI, including normalized difference vegetation index (NDVI) is used to estimate the soybean yield. Combination of various spectral bands, vegetation indices, temperature and estimated soybean yield will be used to classify the different plots using classification and regression trees methods and compared with the manual selection. The findings of this study will facilitate in future breeding line selection program with improved accuracy and consistency.

ABSTRACT. Drought stress is one of the major limiting factors in soybean growth and productivity. Leaf wilting is considered as a common visible symptom of soybeans under drought condition. In soybean breeding programs, genotypes with slow wilting trait have been identified as drought tolerant cultivars. Traditional method uses visual ranking based on experience to discriminate slow-and fast-wilting genotypes, which is subjective and time-consuming. Recent developments in UAV-based high-throughput phenotyping have shown a great potential for measuring plant traits and detecting plant responses under stresses. The goal of this study is to investigate the potential of identifying soybean genotypes with the slow-wilting trait using UAV-based high-throughput phenotyping. A UAV-based platform was developed to collect high-resolution images for large-scale soybean genotypes planted at two locations under drought stress. Spatial analysis is performed to correct image phenotypic information due to experimental design factors and spatial variance. By associated the genetic means with the traditional visual rankings of wilting, a classification tree is performed to discriminate genotypes with different wilting performance. The broad sense heritability (H²) of the significant traits are computed for quantifying the repeatability of the wilting discrimination. The classification model is expected to discriminate soybean leaf wilting at the early stage (R2) in order to improve the efficiency of breeding programs.

ABSTRACT. Although the use of high intensity chemo-radiotherapies have demonstrated only modest improvement in the treatment of aggressive tumors, undesirable drug specific as well as radiation therapy incurred side effects leading up to DNA damage enhances the risk of developing into a second cancer at a later stage, as observed by the scientists and the researchers. An innovative technique gaining traction in the area of cancer therapeutics is novel nanostructures induced hyperthermia of the tumor cells; however, several concerns prevent a biomedical breakthrough.

In this study, an innovative multimodal hyperthermia strategy that has been unexplored thus far – “hybrid photo-magnetic (PMA) stimulation” has successfully been implemented on magnetite (Fe3O4) and gold (Au) nanoparticle (NP) decorated dextran covered carbon nanotube (CNT) structures (DIGCNTs). The designed nanostructures demonstrated excellent biocompatibility; however severely affected the viability of a cellular model in culture when activated by low intensity PMA stimulation. This novel strategy permitted the use of a less intense AC magnetic field in combination with optical irradiation during the treatment, thus removing the safety concerns associated with the AC magnetic field assisted therapies. Additionally, the treatment efficacy has been achieved at a reduced nanoparticle dose level.

Thus, our study suggests that smart nanostructure-based hybrid PMA irradiation is a safer and viable approach to remotely guide cell destruction, which may be adopted as an efficient technique in clinical management to develop in-vivo hyperthermia.

ABSTRACT. Macroporous cell-laden hydrogels have recently gained recognition for a wide range of biomedical and bioengineering applications. There are various approaches to create porosity in hydrogels such as lyophilization or foam formation, to name a few. However, many do not allow a precise control over pore size or are not compatible with in situ cell encapsulation. Here, we developed novel templated macroporous hydrogels by encapsulating uniform degradable hydrogel microspheres produced via microfluidics into a hydrogel slab. The microspheres degraded completely leaving macropores behind. Microsphere degradation was dependent on the incubation medium, microspheres size, microsphere confinement in the hydrogel as well as cell encapsulation. Uniquely, the degradable microspheres were biocompatible and when laden with glioblastoma cells, the cells were deposited in the macropores upon microsphere degradation and formed multicellular aggregates. The hydrogel-encapsulated glioblastoma aggregates were used in a small drug screen to demonstrate the relevance of cell-matrix interactions for multicellular spheroid drug responsiveness. Hydrogel-grown spheroid cultures are increasingly important in applications such as in vitro tumor, hepatocellular, and neurosphere cultures and drug screening; hence, the templated cell aggregate-laden hydrogels described here would find utility in various applications.

ABSTRACT. Zeolites are a group of nanostructured, microporous materials that have been suggested as a possible material for biomedical applications in implantable device coatings, tissue engineering, and drug delivery systems due to their unique interactions with biomolecules and biological environments. Here we demonstrate the fundamental sorption interactions between a pure-silica zeolite (MFI) with lysozyme using enzyme-linked immunosorbent assays (ELISAs) and bicinchoninic acid assays (BCAs). The sorption of lysozyme onto / into pure-silica zeolite MFI was measured against three sets of parameters: the orientation of the thin film’s crystal structure (b-oriented or randomly-oriented), the lysozyme incubation volume (200 µL, 400 µL, 600 µL, 800 µL), and the lysozyme incubation time (1, 6, and 24 hours). The resulting data demonstrates that the films’ ability to sorb lysozyme depends strongly on the specific uptake parameters, make pure-silica zeolite MFI thin films a tailorable candidate for drug delivery and tissue engineering applications.