Implantable Sensors for Regenerative Medicine.

The translation of many tissue engineering/regenerative medicine (TE/RM) therapies that demonstrate promise in vitro are delayed or abandoned due to reduced and inconsistent efficacy when implemented in more complex and clinically relevant preclinical in vivo models. Determining mechanistic reasons for impaired treatment efficacy is challenging after a regenerative therapy is implanted due to technical limitations in longitudinally measuring the progression of key environmental cues in vivo. The ability to acquire real-time measurements of environmental parameters of interest including strain, pressure, pH, temperature, oxygen tension, and specific biomarkers within the regenerative niche in situ would significantly enhance the information available to tissue engineers to monitor and evaluate mechanisms of functional healing or lack thereof. Continued advancements in material and fabrication technologies utilized by microelectromechanical systems (MEMSs) and the unique physical characteristics of passive magnetoelastic sensor platforms have created an opportunity to implant small, flexible, low-power sensors into preclinical in vivo models, and quantitatively measure environmental cues throughout healing. In this perspective article, we discuss the need for longitudinal measurements in TE/RM research, technical progress in MEMS and magnetoelastic approaches to implantable sensors, the potential application of implantable sensors to benefit preclinical TE/RM research, and the future directions of collaborative efforts at the intersection of these two important fields.

Wireless Implantable Sensor for Noninvasive, Longitudinal Quantification of Axial Strain Across Rodent Long Bone Defects.

Bone development, maintenance, and regeneration are remarkably sensitive to mechanical cues. Consequently, mechanical stimulation has long been sought as a putative target to promote endogenous healing after fracture. Given the transient nature of bone repair, tissue-level mechanical cues evolve rapidly over time after injury and are challenging to measure noninvasively. The objective of this work was to develop and characterize an implantable strain sensor for noninvasive monitoring of axial strain across a rodent femoral defect during functional activity. Herein, we present the design, characterization, and in vivo demonstration of the device's capabilities for quantitatively interrogating physiological dynamic strains during bone regeneration. Ex vivo experimental characterization of the device showed that it possessed promising sensitivity, signal resolution, and electromechanical stability for in vivo applications. The digital telemetry minimized power consumption, enabling extended intermittent data collection. Devices were implanted in a rat 6 mm femoral segmental defect model, and after three days, data were acquired wirelessly during ambulation and synchronized to corresponding radiographic videos, validating the ability of the sensor to noninvasively measure strain in real-time. Together, these data indicate the sensor is a promising technology to quantify tissue mechanics in a specimen specific manner, facilitating more detailed investigations into the role of the mechanical environment in dynamic bone healing and remodeling processes.

Sensing nitrous oxide with QCL-coupled silicon-on-sapphire ring resonators.

We report the initial evaluation of a mid-infrared QCL-coupled silicon-on-sapphire ring resonator gas sensor. The device probes the N(2)O 2241.79 cm(-1) optical transition (R23 line) in the ν(3) vibrational band. N(2)O concentration is deduced using a non-linear least squares fit, based on coupled-mode theory, of the change in ring resonator Q due to gas absorption losses in the evanescent portion of the waveguide optical mode. These early experiments demonstrated response to 5000 ppmv N(2)O.

Rapid homogeneous endothelialization of high aspect ratio microvascular networks.

Microvascularization of an engineered tissue construct is necessary to ensure the nourishment and viability of the hosted cells. Microvascular constructs can be created by seeding the luminal surfaces of microfluidic channel arrays with endothelial cells. However, in a conventional flow-based system, the uniformity of endothelialization of such an engineered microvascular network is constrained by mass transfer of the cells through high length-to-diameter (L/D) aspect ratio microchannels. Moreover, given the inherent limitations of the initial seeding process to generate a uniform cell coating, the large surface-area-to-volume ratio of microfluidic systems demands long culture periods for the formation of confluent cellular microconduits. In this report, we describe the design of polydimethylsiloxane (PDMS) and poly(glycerol sebacate) (PGS) microvascular constructs with reentrant microchannels that facilitates rapid, spatially homogeneous endothelial cell seeding of a high L/D (2 cm/35 µm; > 550:1) aspect ratio microchannels. MEMS technology was employed for the fabrication of a monolithic, elastomeric, reentrant microvascular construct. Isotropic etching and PDMS micromolding yielded a near-cylindrical microvascular channel array. A 'stretch - seed - seal' operation was implemented for uniform incorporation of endothelial cells along the entire microvascular area of the construct yielding endothelialized microvascular networks in less than 24 h. The feasibility of this endothelialization strategy and the uniformity of cellularization were established using confocal microscope imaging.

A BIOCOMPATIBLE GLASS-ENCAPSULATED TRIAXIAL FORCE SENSOR FOR IMPLANTABLE TACTILE SENSING APPLICATIONS.

This paper reports a microfabricated triaxial capacitive force sensor. The sensor is fully encapsulated with inert and biocompatible glass (fused silica) material. The sensor comprises two glass plates, on which four capacitors are located. The sensor is intended for subdermal implantation in fingertips and palms and providing tactile sensing capabilities for patients with paralyzed hands. Additional electronic components, such as passives and IC chips, can also be integrated with the sensor in a hermetic glass package to achieve an implantable tactile sensing system. Through attachment to a human palm, the sensor has been shown to respond appropriately to typical hand actions, such as squeezing or picking up a bottle.

Optimal bilayer composites for temperature-tracking wireless electronics.

Modern silicone-based epidermal electronics engineered for body temperature sensing represent a pivotal development in the quest for advancing preventive medicine and enhancing post-surgical monitoring. While these compact and highly flexible electronics empower real-time monitoring in dynamic environments, a noteworthy limitation is the challenge in regulating the infiltration or obstruction of heat from the external environment into the surface layers of these electronics. The study presents a cost-effective temperature sensing solution by embedding wireless electronics in a multi-layered elastomeric composite to meet the dual needs of enhanced thermal insulation for encapsulation in contact with air and improved thermal conductivity for the substrate in contact with the skin. The encapsulating composite benefits from the inclusion of hollow silica microspheres, which reduce the thermal conductivity by 40%, while non-spherical aluminum nitride enhances the thermal conductivity of the substrate by 370%. The addition of particles to the respective composites inevitably leads to an increase in modulus. Two composite elements are engineered to coexist while maintaining a matching low modulus of 3.4 MPa and a stretchability exceeding 30%, all without compromising the optimized thermal properties. Consecutive thermal, electrical, and mechanical characterization confirms the sensor's capacity for precise body temperature monitoring during a single day's lifespan, while also assessing the influence of behavioral factors on body temperature.

Hollow microneedles for intradermal injection fabricated by sacrificial micromolding and selective electrodeposition.

Limitations with standard intradermal injections have created a clinical need for an alternative, low-cost injection device. In this study, we designed a hollow metal microneedle for reliable intradermal injection and developed a high-throughput micromolding process to produce metal microneedles with complex geometries. To fabricate the microneedles, we laser-ablated a 70 µm × 70 µm square cavity near the tip of poly(lactic acid) (PLA) microneedles. The master structure was a template for multiple micromolded poly(lactic acid-co-glycolic acid) (PLGA) replicas. Each replica was sputtered with a gold seed layer with minimal gold deposited in the cavity due to masking effects. In this way, nickel was electrodeposited selectively outside of the cavity, after which the polymer replica was dissolved to produce a hollow metal microneedle. Force-displacement tests showed the microneedles, with 12 µm thick electrodeposition, could penetrate skin with an insertion force 9 times less than their axial failure force. We injected fluid with the microneedles into pig skin in vitro and hairless guinea pig skin in vivo. The injections targeted 90 % of the material within the skin with minimal leakage onto the skin surface. We conclude that hollow microneedles made by this simple microfabrication method can achieve targeted intradermal injection.

High-Energy-Density Zinc-Air Microbatteries with Lean PVA-KOH-K2CO3 Gel Electrolytes.

Small-scale, primary electrochemical energy storage devices ("microbatteries") are critical power sources for microelectromechanical system (MEMS)-based sensors and actuators. However, the achievable volumetric and gravimetric energy densities of microbatteries are typically insufficient for intermediate-term applications of MEMS-enabled distributed internet-connected devices. Further, in the increasing subset of Internet of Things (IoT) nodes, where actuation is desired, the peak power density of the microbattery must be simultaneously considered. Metal-air approaches to achieving microbatteries are attractive, as the anode and cathode are amenable to miniaturization; however, further improvements in energy density can be obtained by minimizing the electrolyte volume. To investigate these potential improvements, this work studied very lean hydrogel electrolytes based on poly(vinyl alcohol) (PVA). Integration of high potassium hydroxide (KOH) loading into the PVA hydrogel improved electrolyte performance. The addition of potassium carbonate (K2CO3) to the KOH-PVA gel decreased the carbonation consumption rate of KOH in the gel electrolyte by 23.8% compared to PVA-KOH gel alone. To assess gel performance, a microbattery was formed from a zinc (Zn) anode layer, a gel electrolyte layer, and a carbon-platinum (C-Pt) air cathode layer. Volumetric energy densities of approximately 1400 Wh L-1 and areal peak power densities of 139 mW cm-2 were achieved with a PVA-KOH-K2CO3 electrolyte. Further structural optimization, including using multilayer gel electrolytes and thinning the air cathode, resulted in volumetric and gravimetric energy densities of 1576 Wh L-1 and 420 Wh kg-1, respectively. The batteries described in this work are manufactured in an open environment and fabricated using a straightforward layer-by-layer method, enabling the potential for high fabrication throughput in a MEMS-compatible fashion.

An implantable wireless tactile sensing system.

The sense of touch is critical to dexterous use of the hands and thus an essential component to efforts to restore hand function after amputation or paralysis. Prosthetic systems have focused on wearable tactile sensors. But wearable sensors are suboptimal for neuroprosthetic systems designed to reanimate a patient's own paralyzed hand. Here, we developed an implantable tactile sensing system intended for subdermal placement. The system is composed of a microfabricated capacitive force sensor, a custom integrated circuit supporting wireless powering and data transmission, and a laser-fused hermetic silica package. The miniature device was validated through simulations, benchtop testing, and ex vivo testing in a primate hand. The sensor implanted in the fingertip accurately measured skin forces with a resolution of 4.3 mN. The output from this novel sensor could be encoded in the brain with microstimulation to provide tactile feedback. More broadly, the materials, system design, and fabrication approach establish new foundational capabilities for various applications of implantable sensing systems.

An implantable, wireless, battery-free system for tactile pressure sensing.

The sense of touch is critical to dexterous use of the hands and thus an essential component of efforts to restore hand function after amputation or paralysis. Prosthetic systems have addressed this goal with wearable tactile sensors. However, such wearable sensors are suboptimal for neuroprosthetic systems designed to reanimate a patient's own paralyzed hand. Here, we developed an implantable tactile sensing system intended for subdermal placement. The system is composed of a microfabricated capacitive pressure sensor, a custom integrated circuit supporting wireless powering and data transmission, and a laser-fused hermetic silica package. The miniature device was validated through simulations, benchtop assessment, and testing in a primate hand. The sensor implanted in the fingertip accurately measured applied skin forces with a resolution of 4.3 mN. The output from this novel sensor could be encoded in the brain with microstimulation to provide tactile feedback. More broadly, the materials, system design, and fabrication approach establish new foundational capabilities for various applications of implantable sensing systems.

Intracellular protein delivery and gene transfection by electroporation using a microneedle electrode array.

The impact of many biopharmaceuticals, including protein- and gene-based therapies, has been limited by the need for better methods of delivery into cells within tissues. Here, intracellular delivery of molecules and transfection with plasmid DNA by electroporation is presented using a novel microneedle electrode array designed for the targeted treatment of skin and other tissue surfaces. The microneedle array is molded out of polylactic acid. Electrodes and circuitry required for electroporation are applied to the microneedle array surface by a new metal-transfer micromolding method. The microneedle array maintains mechanical integrity after insertion into pig cadaver skin and is able to electroporate human prostate cancer cells in vitro. Quantitative measurements show that increasing electroporation pulse voltage increases uptake efficiency of calcein and bovine serum albumin, whereas increasing pulse length has lesser effects over the range studied. Uptake of molecules by up to 50% of cells and transfection of 12% of cells with a gene for green fluorescent protein is demonstrated at high cell viability. It is concluded that the microneedle electrode array is able to electroporate cells, resulting in intracellular uptake of molecules, and has potential applications to improve intracellular delivery of proteins, DNA, and other biopharmaceuticals.

Highly-compliant, microcable neuroelectrodes fabricated from thin-film gold and PDMS.

Bio-electrodes have traditionally been made of materials such as metal and silicon that are much stiffer than the tissue from which they record or stimulate. This difference in mechanical compliance can cause incomplete or ineffective contact with the tissue. The electrode stiffness has also been hypothesized to cause chronic low-grade injury and scar-tissue encapsulation, reducing stimulation and recording efficiency. As an initial step to resolve these issues with electrode performance, we have developed and characterized electrically-functional, low-Young's modulus, microcable-shaped neuroelectrodes and demonstrated electrophysiological recording functionality. The microcable geometry gives the electrodes a similar footprint to traditional wire and microwire neuroelectrodes, while reducing the difference in Young's modulus from nervous tissue by orders of magnitude. The electrodes are composed of PDMS and thin-film gold, affording them a high-level of compliance that is well suited for in vivo applications. The composite Young's modulus of the electrode was experimentally determined to be 1.81 ± 0.01 MPa. By incorporating a high-tear-strength silicone, Sylgard 186, the load at failure was increased by 92%, relative to that of the commonly used Sylgard 184. The microcable electrodes were also electromechanically tested, with measurable conductivity (220 kΩ) at an average 8% strain (n = 2) after the application of 200% strain. Electrophysiological recording is demonstrated by wrapping the electrode around a peripheral nerve, utilizing the compliance and string-like profile of the electrode for effective recording in nerve tissue.

Inclined nanoimprinting lithography for 3D nanopatterning.

We report a non-conventional shear-force-driven nanofabrication approach, inclined nanoimprint lithography (INIL), for producing 3D nanostructures of varying heights on planar substrates in a single imprinting step. Such 3D nanostructures are fabricated by exploiting polymer anisotropic dewetting where the degree of anisotropy can be controlled by the magnitude of the inclination angle. The feature size is reduced from micron scale of the template to a resultant nanoscale pattern. The underlying INIL mechanism is investigated both experimentally and theoretically. The results indicate that the shear force generated at a non-zero inclination angle induced by the INIL apparatus essentially leads to asymmetry in the polymer flow direction ultimately resulting in 3D nanopatterns with different heights. INIL removes the requirements in conventional nanolithography of either utilizing 3D templates or using multiple lithographic steps. This technique enables various 3D nanoscale devices including angle-resolved photonic and plasmonic crystals to be fabricated.

All-Passive Hardware Implementation of Multilayer Perceptron Classifiers.

Bottom-up-fabricated crossbars promise superior circuit density and 3-D integrability compared with the traditional CMOS-based implementations. However, their inherent stochasticity presents difficulties in building complex circuits from components that demand precise patterning and high registration accuracies. With fewer terminals than active devices, passive components offer higher device densities and registration tolerances, making them amenable to bottom-up synthesized nanocrossbars. Motivated by this preference for passivity, we explore, in this article, neuromorphic classifiers based on passive neurons and passive synapses. We demonstrate via SPICE simulations how a shallow network of the diode-resistor-based passive rectifier neurons and resistive voltage summers, despite its inherent inability to buffer, amplify, and negate signals, can recognize MNIST digits with 95.4% accuracy. We introduce weight-to-conductance mappings that enable negative weights to be implemented in hardware without excessive memory overheads. The influences of soft and hard defects on the classification performance are evaluated, and the methods to boost fault-tolerance are proposed. The first-order evaluation of the area, speed, and power consumption of the passive multilayer perceptron classifiers is undertaken, and the results are compared with a benchmark study in neuromorphic hardware.

A microwell-based impedance sensor on an insertable microneedle for real-time in vivo cytokine detection.

Impedance-based protein detection sensors for point-of-care diagnostics require quantitative specificity, as well as rapid or real-time operation. Furthermore, microfabrication of these sensors can lead to the formation of factors suitable for in vivo operation. Herein, we present microfabricated needle-shaped microwell impedance sensors for rapid-sample-to-answer, label-free detection of cytokines, and other biomarkers. The microneedle form factor allows sensors to be utilized in transcutaneous or transvascular sensing applications. In vitro, experimental characterization confirmed sensor specificity and sensitivity to multiple proteins of interest. Mechanical characterization demonstrated sufficient microneedle robustness for transcutaneous insertion, as well as preserved sensor function postinsertion. We further utilized these sensors to carry out real-time in vivo quantification of human interleukin 8 (hIL8) concentration levels in the blood of transgenic mice that endogenously express hIL8. To assess sensor functionality, hIL8 concentration levels in serum samples from the same mice were quantified by ELISA. Excellent agreement between real-time in vivo sensor readings in blood and subsequent ELISA serum assays was observed over multiple transgenic mice expressing hIL8 concentrations from 62 pg/mL to 539 ng/mL.

Single-step label-free nanowell immunoassay accurately quantifies serum stress hormones within minutes.

A non-faradaic label-free cortisol sensing platform is presented using a nanowell array design, in which the two probe electrodes are integrated within the nanowell structure. Rapid and low volume (≤5 µl) sensing was realized through functionalizing nanoscale volume wells with antibodies and monitoring the real-time binding events. A 28-well plate biochip was built on a glass substrate by sequential deposition, patterning, and etching steps to create a stack nanowell array sensor with an electrode gap of 40 nm. Sensor response for cortisol concentrations between 1 and 15 µg/dl in buffer solution was recorded, and a limit of detection of 0.5 µg/dl was achieved. Last, 65 human serum samples were collected to compare the response from human serum samples with results from the standard enzyme-linked immunosorbent assay (ELISA). These results confirm that nanowell array sensors could be a promising platform for point-of-care testing, where real-time, laboratory-quality diagnostic results are essential.

An electrically active microneedle array for electroporation.

We have designed and fabricated a microneedle array with electrical functionality with the final goal of electroporating skin's epidermal cells to increase their transfection by DNA vaccines. The microneedle array was made of polymethylmethacrylate (PMMA) by micromolding technology from a polydimethylsiloxane (PDMS) mold, followed by metal deposition, patterning using laser ablation, and electrodeposition. This microneedle array possessed sufficient mechanical strength to penetrate human skin in vivo and was also able to electroporate both red blood cells and human prostate cancer cells as an in vitro model to demonstrate cell membrane permeabilization. A computational model to predict the effective volume for electroporation with respect to applied voltages was constructed from finite element simulation. This study demonstrates the mechanical and electrical functionalities of the first MEMS-fabricated microneedle array for electroporation, designed for DNA vaccine delivery.

Off-axis integrated cavity output spectroscopy with a mid-infrared interband cascade laser for real-time breath ethane measurements.

Cavity-enhanced tunable diode laser absorption spectroscopy is an attractive method for measuring small concentrations of gaseous species. Ethane is a breath biomarker of lipid peroxidation initiated by reactive oxygen species. A noninvasive means of quickly quantifying oxidative stress status has the potential for broad clinical application. We present a simple, compact system using off-axis integrated cavity output spectroscopy with an interband cascade laser and demonstrate its use in real-time measurements of breath ethane. We demonstrate a detection sensitivity of 0.48 ppb/Hz(1/2).

Two-color-absorption sensor for time-resolved measurements of gasoline concentration and temperature.

A midinfrared absorption sensor for crank-angle-resolved in-cylinder measurements of gasoline concentration and gas temperature for spark-ignition internal-combustion engines is reported, and design considerations and validation testing in the controlled environments of a heated cell and shock-heated gases are discussed. Mid-IR laser light was tuned to transitions in the strong absorption bands associated with C-H stretching vibration near 3.4 microm, and time-resolved fuel vapor concentration and gas temperature were determined simultaneously from the absorption at two different wavelengths. These two infrared laser wavelengths were simultaneously produced by difference-frequency generation, which combines a near-IR signal laser with two near-IR pump lasers in a periodically poled lithium niobate crystal. Injection current modulation of the pump lasers produced intensity modulation of the mid-IR, which allowed the transmitted signals from the two laser wavelengths to be detected on a single detector and separated by frequency demultiplexing. Injection current modulation produced a wavelength modulation synchronous with the intensity modulation for each of the laser wavelengths, and accurate measurement of the gasoline absorption signal required the effects of wavelength modulation to be considered. Validation experiments were conducted for a single-component hydrocarbon fuel (2,2,4-trimethyl-pentane, commonly known as iso-octane) and a gasoline blend in a heated static cell (300 < or = T < or = 600 K) and behind planar shock waves (600 < T < 1100 K) in a shock tube. With a bandwidth of 10 kHz, the measured fuel concentrations agreed within 5% RMS and the measured temperature agreed within 3% RMS to the known values. The 10 kHz bandwidth is sufficient to resolve 1 crank-angle degree at 1600 RPM.

Electrical Interconnects Fabricated From Biodegradable Conductive Polymer Composites.

This study presents the development and characterization of biodegradable electrical interconnects for transient implantable medical devices. The interconnects comprised micropatterned biodegradable conductive polymer composites, which were developed using iron (Fe) microparticles as the conductive filler and polycaprolactone (PCL) as the insulating matrix. The electrical properties of the composites were investigated under various degradation conditions. Electrical percolation was observed at 17% iron volume fraction, but higher volume fractions exhibited more stable electrical resistivity throughout the time course of physiological degradation. The electrical resistivity of 40%vf Fe-PCL composites increased tenfold in an emulated packaged environment under degradation. Biodegradable electrical interconnects based on 40%vf Fe-PCL composites were successfully micropatterned in daisy chain structures, illustrating the process compatibility of Fe-PCL composites for interconnect applications. The electrical resistance of the packaged daisy chain structures exhibited a reasonable increase under degradation. An electrical lifetime of over 5 days was also achieved. System integration with a commercial humidity sensor and analytical calculations supporting other application scenarios confirmed the feasibility of micropatterned Fe-PCL interconnects for use in implantable electrical systems.