A Novel Vascular Anastomotic Coupling Device for End-to-End Anastomosis of Arteries and Veins.

OBJECTIVE: Hand-sutured (HS) techniques remain the gold standard for most microvascular anastomoses in microsurgery. HS techniques can result in endothelial lacerations and back wall suturing, leading to complications such as thrombosis and free tissue loss. A novel force-interference-fit vascular coupling device (FIF-VCD) system can potentially reduce the need for HS and improve end-to-end anastomosis. This study aims to describe the development and testing of a novel FIF-VCD system for 1.5 to 4.0 mm outside diameter arteries and veins. METHODS: Benchtop anastomoses were performed using porcine cadaver arteries and veins. Decoupling force and anastomotic leakage were tested under simulated worst-case intravital physiological conditions. The 1.5 mm FIF-VCD system was used to perform cadaver rat abdominal aorta anastomoses. RESULTS: Benchtop testing showed that the vessels coupled with the FIF-VCD system could withstand simulated worst-case intravital physiological conditions with a 95% confidence interval for the average decoupling force safety factor of 8.2 ± 1.0 (5.2 ± 1.0 N) and a 95% confidence interval for the average leakage rate safety factor of 26 ± 3.6 (8.4 ± 0.14 and 95 ± 1.4 µL/s at 150 and 360 mmHg, respectively) when compared to HS anastomotic leakage rates (310 ± 14 and 2,100 ± 72 µL/s at 150 and 360 mmHg, respectively). The FIF-VCD system was successful in performing cadaver rat abdominal aorta anastomoses. CONCLUSION: The FIF-VCD system can potentially replace HS in microsurgery, allowing the safe and effective connection of arteries and veins. Further studies are needed to confirm the clinical viability and effectiveness of the FIF-VCD system.

Characterizing a Silver Nanoparticle-Based Electrochemical Biosensor for Shiga Toxin Detection.

Shiga toxins (1, 2) regularly cause outbreaks and food recalls and pose a significant health risk to the infected population. Therefore, new reliable tools are needed to rapidly detect Shiga toxin cost-effectively in food, water, and wastewater before human consumption. Enzyme immunoassay and polymerase chain reaction approaches are the gold standard detection methods for the Shiga toxin. However, these methods require expensive instruments along with expensive reagents, which makes them hard to convert into point-of-use and low-cost systems. This study introduces an electrochemical biosensing method that utilizes silver nanoparticles (AgNPs) as electrochemical tags and commercially available low-cost screen-printed carbon electrodes for detection. This study introduces the modification of reference electrodes on commercially available screen-printed carbon electrodes to detect AgNPs dissolved in nitric acid. This biosensor achieved a 2 ng/mL lowest measured concentration for Shiga toxin-1 in less than 3 h. These biosensor results also showed that the AgNP-based sensor has better linearity (for graph between peak current vs concentration) and lower standard deviation compared to gold nanoparticles (AuNP)-based electrochemical biosensors.

Local delivery of FK506 to a nerve allograft is comparable to systemic delivery at suppressing allogeneic graft rejection.

The objective of this study was to determine if locally delivered FK506 could prevent allogeneic nerve graft rejection long enough to allow axon regeneration to pass through the nerve graft. An 8mm mouse sciatic nerve gap injury repaired with a nerve allograft was used to assess the effectiveness of local FK506 immunosuppressive therapy. FK506-loaded poly(lactide-co-caprolactone) nerve conduits were used to provide sustained local FK506 delivery to nerve allografts. Continuous and temporary systemic FK506 therapy to nerve allografts, and autograft repair were used as control groups. Serial assessment of inflammatory cell and CD4+ cell infiltration into the nerve graft tissue was performed to characterize the immune response over time. Nerve regeneration and functional recovery was serially assessed by nerve histomorphometry, gastrocnemius muscle mass recovery, and the ladder rung skilled locomotion assay. At the end of the study, week 16, all the groups had similar levels of inflammatory cell infiltration. The local FK506 and continuous systemic FK506 groups had similar levels of CD4+ cell infiltration, however, it was significantly greater than the autograft control. In terms of nerve histmorphometry, the local FK506 and continunous systemic FK506 groups had similar amounts of myelinated axons, although they were significantly lower than the autograft and temporary systemic FK506 group. The autograft had significantly greater muscle mass recovery than all the other groups. In the ladder rung assay, the autograft, local FK506, and continuous systemic FK506 had similar levels of skilled locomotion performance, whereas the temporary systemic FK506 group had significanty better performance than all the other groups. The results of this study suggest that local delivery of FK506 can provide comparable immunosuppression and nerve regeneration outcomes as systemically delivered FK506.

Evaluating the influence of particle morphology and density on the viscosity and injectability of a novel long-acting local anesthetic suspension.

Proper pain management is well understood to be one of the fundamental aspects of a healthy postoperative recovery in conjunction with mobility and nutrition. Approximately, 10% of patients prescribed opioids after surgery continue to use opioids in the long-term and as little as 10 days on opioids can result in addiction. In an effort to provide physicians with an alternative pain management technique, this work evaluates the material properties of a novel local anesthetic delivery system designed for controlled release of bupivacaine for 72 hours. The formulation utilizes solid-lipid microparticles that encapsulate the hydrophobic molecule bupivacaine in its free-base form. The lipid microparticles are suspended in a non-crosslinked hyaluronic acid hydrogel, which acts as the microparticle carrier. Two different particle manufacturing techniques, milling and hot homogenization, were evaluated in this work. The hot homogenized particles had a slower and more controlled release than the milled particles. Rheological techniques revealed that the suspension remains a viscoelastic fluid when loaded with either particle type up to 25% (w/v) particles densities. Furthermore, the shear thinning properties of the suspension media, hyaluronic acid hydrogel, were conserved when bupivacaine-loaded solid-lipid microparticles were loaded up to densities of 25% (w/v) particle loading. The force during injection was measured for suspension formulations with varying hyaluronic acid hydrogel concentrations, particle densities, particle types and particle sizes. The results indicate that the formulation viscosity is highly dependent on particle density, but hyaluronic acid hydrogel is required for lowering injection forces as well as minimizing clogging events.

Separation of U87 glioblastoma cell-derived small and medium extracellular vesicles using elasto-inertial flow focusing (a spiral channel).

Nanoscale and microscale cell-derived extracellular vesicle types and subtypes are of significant interest to researchers in biology and medicine. Extracellular vesicles (EVs) have diagnostic and therapeutic potential in terms of biomarker and nanomedicine applications. To enable such applications, EVs must be isolated from biological fluids or separated from other EV types. Developing methods to fractionate EVs is of great importance to EV researchers. Our goal was to begin to develop a device that would separate medium EVs (mEVs, traditionally termed microvesicles or shedding vesicles) and small EVs (sEVs, traditionally termed exosomes) by elasto-inertial effect. We sought to develop a miniaturized technology that works similar to and provides the benefits of differential ultracentrifugation but is more suitable for EV-based microfluidic applications. The aim of this study was to determine whether we could use elasto-inertial focusing to re-isolate and recover U87 mEVs and sEVs from a mixture of mEVs and sEVs isolated initially by one round of differential ultracentrifugation. The studied spiral channel device can continuously process 5 ml of sample fluid per hour. Using the channel, sEVs and mEVs were recovered and re-isolated from a mixture of U87 glioma cell-derived mEVs and sEVs pre-isolated by one round of differential ultracentrifugation. Following two passes through the spiral channel, approximately 55% of sEVs were recovered with 6% contamination by mEVs (the recovered sEVs contained 6% of the total mEVs). In contrast, recovery of U87 mEVs and sEVs re-isolated using a typical second centrifugation wash step was only 8% and 53%, respectively. The spiral channel also performed similar to differential ultracentrifugation in reisolating sEVs while significantly improving mEV reisolation from a mixture of U87 sEVs and mEVs. Ultimately this technology can also be coupled to other microfluidic EV isolation methods in series and/or parallel to improve isolation and minimize loss of EV subtypes.

Experiment, theory, and simulation of a flow-electrical-split flow thin particle separation device.

Herein, we describe the simulation of a novel flow-electrical-split flow thin (Fl-El-SPLITT) separation device and validate it using existing theory and experimentation for the first time using polystyrene particles of 28 and 1000 nm diameters. The fraction of particles exiting selected ports with DC El-SPLITT is predicted with existing theory, but the theory does not include AC fields, nor does it incorporate the use of crossflows. Using DC fields the El-SPLITT simulation and theory calculated transition points result in the same values. These calculated values accurately predict the experimentally obtained transition point using a 50:50 outlet splitting plane (OSP). Relative to actual experimentally obtained transition points, the calculated values lag behind for a 90:10 OSP, and lead ahead for a 10:90 OSP. The simulation explains trends seen in AC testing, and reasonably predicts the fraction of particles exiting each port. As DC current increases, the amount of AC current required to scatter the particles away from the DC-intended port decreases. The simulation also models a crossflow in a SPLITT system with a DC current applied in a direction opposite the crossflow with some success. Long term steady-state testing without crossflows shows a DC voltage dependent loss of particles. At 8 V DC, total recovery of 28 and 1000 nm particles was 70% and 26%, respectively. This work effectively models a new Fl-El-SPLITT system via Matlab simulation by demonstrating key experimental results such as the influence of DC, AC, and crossflows on the SPLITT separation of polystyrene particles.

Compression of the vascular wall to create a friction fit in a vascular anastomotic coupler.

A previously reported microvascular coupler was shown to effectively create vascular anastomoses, but was too large for practical clinical use. To safely reduce coupler size, certain failure modes needed to be better understood. The coupler functions, in part, by compressing the vessel wall between two concentric rings, creating a friction fit that anchors the device to the vessel. This work investigates the relationship between vessel wall compression and resulting friction fit strength to ensure reducing coupler size will not unduly increase the risk that this friction fit might fail. Vascular walls were compressed to a specified strain and the tensile force required to overcome the resulting friction was measured. Experiments were conducted with various vessel types (Porcine common carotid artery, splenic artery, and jugular vein), across a range of compressive strains (55-95%), and by using either PEEK or HDPE to compress the vessel. Tensile force was increased at a rate of 5 g/min or held constant for 24 h. For experiments with incrementally increasing force, the force at failure varied with compressive strain via a power function. At 70% compression, PEEK produced 4.6 times stronger friction fits than HDPE, and common carotid arteries and splenic arteries produced 1.8 and 1.3 times stronger fits than jugular veins respectively. For experiments where tensile force was applied for 24 h, much lower forces were required to overcome friction. These results were compared to friction fit failure in a coupler prototype and it was found that the prototypes failed at just 30% of the force required to cause vessel slip under the other test conditions. These results were used to develop a model that predicts the probability of device failure via vessel slipping (one design, smaller than previously reported, was estimated to fail at maximum in vivo axial stress once in 500 anastomoses, a potentially safe level of risk).

Development and Testing of a Continuous Flow-Electrical-Split-Flow Lateral Transport Thin Separation System (Fl-El-SPLITT).

In this work, a new high-volume, continuous particle separation device that separates based upon size and charge is described. Two continuous flow-electrical-split-flow lateral transport thin (Fl-El-SPLITT) device architectures (a platinum electrode on a porous membrane and a porous graphite electrode under a membrane) were developed and shown to improve particle separations over a purely electrical-SPLITT device. The graphite FL-El-SPLITT device architecture achieved the best separation of approximately 60% of small (28 nm) vs large (1000 nm) polystyrene particles. Fl-El-SPLITT (platinum) achieved a 75% separation on a single pass using these same particles. Fl-El-SPLITT (platinum) achieved a moderate 26% continuous separation of U87 glioma cell-derived small extracellular vesicles (EVs) from medium EVs. Control parameter testing showed that El-SPLITT continuously directed particle motility within a channel to exit a selected port based upon the applied voltage using either direct current or alternating current. The transition from one port to the other was dependent upon the voltage applied. Both large and small polystyrene particles transitioned together rather than separating at each of the applied voltages. These data present the first ever validation of El-SPLITT in continuous versus batch format. The Fl-El-SPLITT device architecture, monitoring, and electrical and fluid interfacing systems are described in detail for the first time. Capabilities afforded to the system by the flow addition include enhanced particle separation as well as the ability to filter out small particles or desalinate fluids. High-throughput continuous separations based upon electrophoretic mobility will be streamlined by this new technique that combines electrical and flow fields into a single device.

SARS-CoV-2 pandemic: a review of molecular diagnostic tools including sample collection and commercial response with associated advantages and limitations.

The unprecedented global pandemic known as SARS-CoV-2 has exercised to its limits nearly all aspects of modern viral diagnostics. In doing so, it has illuminated both the advantages and limitations of current technologies. Tremendous effort has been put forth to expand our capacity to diagnose this deadly virus. In this work, we put forth key observations in the functionality of current methods for SARS-CoV-2 diagnostic testing. These methods include nucleic acid amplification-, CRISPR-, sequencing-, antigen-, and antibody-based detection methods. Additionally, we include analysis of equally critical aspects of COVID-19 diagnostics, including sample collection and preparation, testing models, and commercial response. We emphasize the integrated nature of assays, wherein issues in sample collection and preparation could impact the overall performance in a clinical setting.

Modeling diffusion-based drug release inside a nerve conduit in vitro and in vivo validation study.

The objective of this work was to develop a model and understand the diffusion of a drug into and throughout a drug delivering nerve conduit from a surrounding reservoir through a hole in the wall separating the lumen of the conduit and the reservoir. A mathematical model based on Fick's law of diffusion was developed using the finite difference method to understand the drug diffusion and the effect of varying device parameters on the concentration of drug delivered from a hole-based drug delivery device. The mathematical model was verified using a physical microfluidic (µFD) model and an in vitro/in vivo release test using prototype devices. The results of the mathematical model evaluation and microfluidic device testing offered positive insight into the reliability and function of the reservoir and hole-based drug delivering nerve conduit. The mathematical model demonstrated how changing device parameters would change the drug concentration inside the device. It was observed that the drug release in the conduit could be tuned by both concentration scaling and changing the hole size or number of holes. Based on the results obtained from the microfluidic device, the error in the mathematical drug release model was shown to be less than 10% when comparing the data obtained from mathematical model and µFD model. The data highlights the flexibility of having a hole-based drug delivery system, since the drug release can be scaled predictably by changing the device parameters or the concentration of the drug in the reservoir. Graphical abstract .

Size and shape based chromosome separation in the inertial focusing device.

In this paper, we use a spiral channel inertial focusing device for isolation and purification of chromosomes, which are highly asymmetric. The method developed is proposed as a sample preparation process for transchromosomic research. The proposed microfluidics-based chromosome separation approach enables rapid, label-free isolation of bioactive chromosomes and is compatible with chromosome buffer. As part of this work, particle force analysis during the separation process is performed utilizing mathematic models to estimate the expected behavior of chromosomes in the channel and the model validated with experiments employing fluorescent beads. The chromosome sample is further divided into subtypes utilizing fluorescent activated cell sorting , including small condensed chromosomes, single chromosomes, and groups of two chromosomes (four sister chromatids). The separation of chromosome subtypes is realized based on their shape differences in the spiral channel device under high flow rate conditions. When chromosomes become aligned in the shear flow, the balance between the inertial focusing force and the Dean flow drag force is determined by the chromosome projection area and aspect ratio, or shape difference, leading to different focusing locations in the channel. The achieved results indicate a new separation regime in inertial microfluidics that can be used for the separation of non-spherical particles based on particle aspect ratios, which could potentially be applied in fields such as bacteria subtype separation and chromosome karyotyping.

Entrapping bupivacaine-loaded emulsions in a crosslinked-hydrogel increases anesthetic effect and duration in a rat sciatic nerve block model.

The purpose of this research was to evaluate a novel long-acting bupivacaine delivery system for control of postoperative pain. Bupivacaine-loaded lipid emulsion (BLE) droplets were created by high-speed homogenization. The BLE droplets were then entrapped into a crosslinked-hyaluronic acid hydrogel system to create an injectable composite gel formulation (HA-BLE). Dynamic light scattering, rheological, and drug release techniques were used to characterize the formulations. A rat sciatic nerve block with a thermal nociceptive assay was used to evaluate the anesthetic effect in comparison to controls, bupivacaine HCl and liposomal bupivacaine. The BLE droplets had a zeta potential, droplet size, and polydispersity index of -40.8 ± 0.66 mV, 299 ± 1.77 nm, and 0.409 ± 0.037, respectively. The HA-BLE formulation could be injected through 25 g needles and had an elastic modulus of 372 ± 23.7 Pa. Approximately 80% and 100% of bupivacaine was released from the BLE and HA-BLE formulations by 20 and 68 h, respectively. The HA-BLE formulation had a 5-times greater anesthetic area under the curve and an anesthetic duration that was twice as long as controls. Results indicate that incorporating the BLEs into the hydrogel significantly increased anesthetic effect by protecting the BLE droplets from the in vivo environment.

Characterization of Human Glioblastoma versus Normal Plasma-Derived Extracellular Vesicles Preisolated by Differential Centrifugation Using Cyclical Electrical Field-Flow Fractionation.

Although many properties for small extracellular vesicles (sEVs, formerly termed "exosomes") isolated at ∼100 000g are known, a wide range of values are reported for their electrophoretic mobility (EM) measurements. This paper reports for the first time the effect of dilution on the EM of U87 glioblastoma cell-derived and plasma-derived sEVs and medium size EVs (mEVs, commonly termed "oncosomes") preisolated by differential centrifugation. Furthermore, the effect of resalting on the EM of sEVs and mEVs was evaluated. The EM of U87 sEVs and U87 mEVs showed an increase as the salt concentration decreased to 0.005% of the initial salt concentration. However, for the plasma sEVs and plasma mEVs, the electrophoretic mobility increased as the salt concentration decreased to 0.01% of the initial salt concentration and then increased to its initial value when the salt concentration decreased to 0.005% of the initial salt concentration. For both U87 and plasma sEVs and mEVs, the EM remained almost constant when the concentration of the particles changed and the salt concentration was kept the same as its initial value. This indicates that the EM of EVs is only a function of the salt concentration of the buffer and is independent of the concentration of the particles. The sEVs and mEVs were separated with cyclical ElFFF for the first time. The results indicate that ElFFF was able to fractionate the EVs, and a crescent-shaped trend was found for the retention time when the applied AC voltage was altered (increased).

Enhanced chromosome extraction from cells using a pinched flow microfluidic device.

Extraction and purification of intact chromosomes are critical sample preparation steps for transchromosomic research and other applications. The commonly used sample preparation methods lead to too few chromosomes with chromosome deactivation and degradation. In this paper, a "mild" chromosome extraction process that combines a chemical and mechanical lysis approach is introduced for the preparation of intact chromosomes that can readily be used for downstream processing. Metaphase cells are treated by chemical lysis buffer and pushed through a microfluidic pinched flow device. Cells are ruptured, and chromosomes are released by a combination of shear stress and chemical reagents. Chromosomes are released intact from the cell membrane into the solution. Simulations and experiments are performed to optimize the microfluidic device geometry and operation parameters. Cell rupture and chromosome release are found to be improved by the shear stress in the pinched flow device. Simulation results indicate that the maximum shear stress appears in the channel constriction region, and the narrow channel maintains constant shear stress. It is concluded that the constriction design, narrow channel width, and operation flow rate have a significate influence on chromosome release. Utilizing an optimized device, near-complete cell lysis is achieved and 4 times as many chromosomes are released (8% in control experiments to 25% in optimized pinched flow devices). Sample treatment time can also be reduced utilizing this combined chemical-mechanical chromosome release method.

Local FK506 delivery at the direct nerve repair site improves nerve regeneration.

INTRODUCTION: The objective of this study is to assess the efficacy of local tacrolimus (FK506) delivery to improve outcomes in the setting of nerve transection injury. METHODS: FK506 embedded poly(lactide-co-caprolactone) films capable of extended, localized release of FK506 were developed. FK506 rate of release testing and bioactivity assay was performed. Mouse sciatic nerve transection and direct repair model was used to evaluate the effect extended, local delivery of FK506 had on nerve regeneration outcomes. RESULTS: Linear release of FK506 was observed for 30 days and released FK506 matched control levels of neurite extension in the dorsal root ganglion assay. Groups treated with local FK506 had greater gastrocnemius muscle weight, foot electromyogram, and number of axons distal of the repair site than non-FK506 groups. DISCUSSION: Results of this study indicate that extended, localized delivery of FK506 to nerve injuries can improve nerve regeneration outcomes in a mouse sciatic nerve transection and repair.

Hydrodynamic cavitation for the rapid separation and electrochemical detection of Cryptosporidium parvum and Escherichia coli O157:H7 in ground beef.

Foodborne illnesses are a major contributor to misery and health challenges in both rich and poor nations. Illnesses from pathogens such as Escherichia coli and Cryptosporidium parvum oocysts account for most of the cases of diarrhea in the world. Many standard methods exist for detecting these pathogens in water. However, these standard methods do not readily translate to the detection of the same pathogens in food. Detection techniques for pathogens in food are often inadequate, due to their inability to completely separate pathogens from food matrices. In this paper, we present a technique to separate and detect both Escherichia coli cells and Cryptosporidium parvum oocysts that have been embedded in ground meat. We achieve this objective by combining enzymatic digestion of the meat, hydrodynamic cavitation to disassemble pathogens from the meat, immunomagnetic separation to purify meat samples and indirect electrochemical detection of the target pathogens. Our use of hydrodynamic cavitation to separate pathogens is compared against an industry standard separation technique. Results indicate that the use of hydrodynamic cavitation amplifies the detection capabilities of our sensing technique and is overall comparable to or better than conventional stomacher sample preparation.

Drug-delivering nerve conduit improves regeneration in a critical-sized gap.

Autologous nerve grafts are the current "gold standard" for repairing large nerve gaps. However, they cause morbidity at the donor nerve site and only a limited amount of nerve can be harvested. Nerve conduits are a promising alternative to autografts and can act as guidance cues for the regenerating axons, without the need to harvest donor nerve. Separately, it has been shown that localized delivery of GDNF can enhance axon growth and motor recovery. FK506, an FDA approved small molecule, has also been shown to enhance peripheral nerve regeneration. This paper describes the design of a novel hole-based drug delivery apparatus integrated with a polytetrafluoroethylene (PTFE) nerve conduit for controlled local delivery of a protein such as GDNF or a small molecule such as FK506. The PTFE devices were tested in a diffusion chamber, and the bioactivity of the released media was evaluated by measuring neurite growth of dorsal root ganglions (DRGs) exposed to the released drugs. The drug delivering nerve guide was able to release bioactive concentrations of FK506 or GDNF. Following these tests, optimized drug releasing nerve conduits were implanted across 10 mm sciatic nerve gaps in a BL6 yellow fluorescent protein (YFP) mouse model, where they demonstrated significant improvement in muscle mass, compound muscle action potential, and axon myelination in vivo as compared with nerve conduits without the drug. The drug delivery nerve guide could release drug for extended periods of time and enhance axon growth in vitro and in vivo.

Exosome Isolation: Cyclical Electrical Field Flow Fractionation in Low-Ionic-Strength Fluids.

The influence of buffer substitution and dilution effects on exosome size and electrophoretic mobility were shown for the first time. Cyclical electrical field flow fractionation (Cy-El-FFF) in various substituted fluids was applied to exosomes and other particles. Tested carrier fluids of deionized (DI) water, 1× phosphate buffered saline (PBS), 0.308 M trehalose, and 2% isopropyl alcohol (IPA) influenced Cy-El-FFF-mediated isolation of A375 melanoma exosomes. All fractograms revealed a crescent-shaped trend in retention times with increasing voltage with the maximum retention time at ∼1.3 V AC. A375 melanoma exosome recovery was approximately 70-80% after each buffer substitution, and recovery was independent of whether the sample was substituted into 1× PBS or DI water. Exosome dilution in deionized water produced a U-shaped dependence on electrophoretic mobility. The effect of dilution using 1× PBS buffer revealed a very gradual change in electrophoretic mobility of exosomes from ∼-1.6 to -0.1 µm cm/s V, as exosome concentration was decreased. This differed from the use of DI water, where a large change from ∼-5.5 to -0.1 µm cm/s V over the same dilution range was observed. Fractograms of separated A375 melanoma exosomes in two substituted low-ionic-strength buffers were compared with synthetic particle fractograms. Overall, the ability of Cy-El-FFF to separate exosomes based on their size and charge is a highly promising, label-free approach to initially catalogue and purify exosome subtypes for biobanking as well as to enable further exosome subtype interrogations.

Controlled release of FK506 from micropatterned PLGA films: potential for application in peripheral nerve repair.

After decades of research, peripheral nerve injury and repair still frequently results in paralysis, chronic pain and neuropathies leading to severe disability in patients. Current clinically available nerve conduits only provide crude guidance of regenerating axons across nerve gap without additional functionality. FK506 (Tacrolimus), an FDA approved immunosuppressant, has been shown to enhance peripheral nerve regeneration but carries harsh side-effects when delivered systemically. The objective of this study was to develop and evaluate a bioresorbable drug delivery system capable of local extended delivery of FK506 that also provides topological guidance cues to guide axon growth via microgrooves. Photolithography was used to create micropatterned poly(lactide-co-glycolic acid) (PLGA) films embedded with FK506. Non-patterned, 10/10 µm (ridge/groove width), and 30/30 µm patterned films loaded with 0, 1, and 3 µg/cm2 FK506 were manufactured and characterized. In vitro FK506 rate of release testing indicated that the films are capable of an extended (at least 56 days), controlled, and scalable release of FK506. Neurite extension bioactivity assay indicated that FK506 released from the films (concentration of samples tested ranged between 8.46-19.7 ng/mL) maintained its neural bioactivity and promoted neurite extension similar to control FK506 dosages (10 ng/mL FK506). The multi-functional FK506 embedded, micropatterned poly(lactide-co-glycolic acid) films developed in this study have potential to be used in the construction of peripheral nerve repair devices.

Electrochemical Detection of E. coli O157:H7 in Water after Electrocatalytic and Ultraviolet Treatments Using a Polyguanine-Labeled Secondary Bead Sensor.

The availability of clean drinking water is a significant problem worldwide. Many technologies exist for purifying drinking water, however, many of these methods require chemicals or use simple methods, such as boiling and filtering, which may or may not be effective in removing waterborne pathogens. Present methods for detecting pathogens in point-of-use (POU) sterilized water are typically time prohibitive or have limited ability differentiating between active and inactive cells. This work describes a rapid electrochemical sensor to differentially detect the presence of active Escherichia coli (E. coli) O157:H7 in samples that have been partially or completely sterilized using a new POU electrocatalytic water purification technology based on superradicals generated by defect laden titania (TiO2) nanotubes. The sensor was also used to detect pathogens sterilized by UV-C radiation for a comparison of different modes of cell death. The sensor utilizes immunomagnetic bead separation to isolate active bacteria by forming a sandwich assay comprised of antibody functionalized secondary magnetic beads, E. coli O157:H7, and polyguanine (polyG) oligonucleotide functionalized secondary polystyrene beads as an electrochemical tag. The assay is formed by the attachment of antibodies to active receptors on the membrane of E. coli, allowing the sensor to differentially detect viable cells. Ultravioloet (UV)-C radiation and an electrocatalytic reactor (ER) with integrated defect-laden titania nanotubes were used to examine the sensors’ performance in detecting sterilized cells under different modes of cell death. Plate counts and flow cytometry were used to quantify disinfection efficacy and cell damage. It was found that the ER treatments shredded the bacteria into multiple fragments, while UV-C treatments inactivated the bacteria but left the cell membrane mostly intact.