Design and in vitro studies of a needle-type glucose sensor for subcutaneous monitoring.

A new miniaturized glucose oxidase based needle-type glucose microsensor has been developed for subcutaneous glucose monitoring. The sensor is equivalent in shape and size to a 26-guage needle (0.45-mm o.d.) and can be implanted with ease without any incision. The novel configuration greatly facilitates the deposition of enzyme and polymer films so that sensors with characteristics suitable for in vivo use (upper limit of linear range greater than 15 mM, response time less than 5 min, and sensitivity yielding a 5:1 signal-to-background ratio at normal basal glucose levels) can be prepared in high yield (greater than 60%). The sensor response is largely independent of oxygen tension in the normal physiological range. It also exhibits good selectivity against common interferences except for the exogenous drug acetaminophen.     

Simultaneous electrochemical detection of nitric oxide and carbon monoxide generated from mouse kidney organ tissues

A planar-type amperometric dual microsensor for simultaneous detection of nitric oxide and carbon monoxide is presented. The sensor consists of a dual platinum microdisk-based working electrode (WE) and a Ag/AgCl counter/reference electrode covered with an expanded poly(tetrafluoroethylene) (Tetra-tex) gas-permeable membrane. The dual WE possesses two different platinized platinum disks (WE1 and WE2, 250 and 25 microm in diameter, respectively). The larger WE1 is further modified with electrochemical deposition of tin. Use of two sensing disks different in their size as well as in their surface modification produces apparently different sensitivity ratios of NO to CO at WE1 and at WE2 (approximately 2 and approximately 10, respectively) that are induced by favorable CO oxidation on the surface of tin versus platinum. Anodic currents independently measured at WE1 and at WE2 are successfully converted to the concentrations of NO and CO in the co-presence of these gases using the differentiated sensitivities at each electrode. The sensor is evaluated in terms of its analytical performance: respectable linear dynamic range (sub nM to microM); low detection limit (approximately 1 nM for NO and <5 nM for CO); selectivity (over nitrite up to approximately 1 mM); and sensitivity (sufficient for analyzing physiological levels of NO and CO). Using the NO/CO dual microsensor, real-time, simultaneous, direct, and quantitative measurements of NO and CO generated from living biological tissue (mouse, c57, kidney) surfaces, for the first time, are reported.     

Genetic engineering of an allosterically based glucose indicator protein for continuous glucose monitoring by fluorescence resonance energy transfer

Real-time monitoring of blood glucose could vastly reduce a number of the long-term complications associated with diabetes. In this article, we present a novel approach that relies on a glucose-binding protein engineered such that a 20% reduction in fluorescence due to the fluorescence resonance energy transfer occurs as a result of glucose binding. This change in fluorescence provides a signal for the optical detection of glucose. The novel glucose indicator protein (GIP) was created by fusing two fluorescent reporter proteins (green fluorescent proteins) to each end of an Escherichia coli glucose-binding protein in such a manner that the spatial separation between the fluorescent moieties changes when glucose binds, thus generating a distinct optical signal that can be used for glucose detection. By placing the GIP within a dialysis hollow fiber sensor, a microsensor has been developed for continuous monitoring of glucose. The sensor had a response time to sudden glucose changes within 100 s and was reversible. The sensor was shown to have an optional range on the order of 10 microM of glucose.     

A microscale biosensor for methane containing methanotrophic bacteria and an internal oxygen reservoir

A microscale biosensor for continuous measurement of methane partial pressure based on a novel counterdiffusion principle is presented. Methane-oxidizing bacteria placed in the microsensor utilize oxygen from an internal oxygen reservoir when methane from the exterior diffuses through the tip membrane. The transducer is an internal oxygen microsensor with its tip positioned between the oxygen reservoir and the sensor tip membrane. The external partial pressure of methane determines the rate of bacterial oxygen consumption within the sensor, which in turn is reflected by the signal from the transducer. Tip diameters were down to 20 μm, enabling us to study methane distribution on a microscale. The microscale construction also results in a low stirring sensitivity and a 95% response time down to 20 s. By tailoring the geometry, sensors can be made to exhibit a linear response in the full range of 0-1 atm partial pressure of methane or, alternatively, to exhibit a linear response only at lower concentrations, improving the sensitivity to below 0.1 kPa, corresponding to ～1 μM in aqueous solution. Temperature, oxygen, and H(2)S interfere with the signal; no interferences were detected from H(2), NH(3), CO(2), or acetate.     

Development and application of a self-referencing glucose microsensor for the measurement of glucose consumption by pancreatic beta-cells

Glucose gradients generated by an artificial source and beta-cells were measured using an enzyme-based glucose microsensor, 8-microm tip diameter, as a self-referencing electrode. The technique is based on a difference measurement between two locations in a gradient and thus allows us to obtain real-time flux values with minimal impact of sensor drift or noise. Flux values were derived by incorporation of the measured differential current into Fick's first equation. In an artificial glucose gradient, a flux detection limit of 8.2 +/- 0.4 pmol.cm(-2).s(-1) (mean +/- SEM, n = 7) with a sensor sensitivity of 7.0 +/- 0.4 pA/ mM (mean +/- SEM, n = 16) was demonstrated. Under biological conditions, the glucose sensor showed no oxygen dependence with 5 mM glucose in the bulk medium. The addition of catalase to the bulk medium was shown to ameliorate surface-dependent flux distortion close to specimens, suggesting an underlying local accumulation of hydrogen peroxide. Glucose flux from beta-cell clusters, measured in the presence of 5 mM glucose, was 61.7 +/- 9.5 fmol.nL(-1).s(-1) (mean +/- SEM, n = 9) and could be pharmacologically modulated. Glucose consumption in response to FCCP (1 microM) transiently increased, subsequently decreasing to below basal by 93 +/- 16 and 56 +/- 6%, respectively (mean +/- SEM, n = 5). Consumption was decreased after the application of 10 microM rotenone by 74 +/- 5% (mean +/- SEM, n = 4). These results demonstrate that an enzyme-based amperometric microsensor can be applied in the self-referencing mode. Further, in obtaining glucose flux measurements from small clusters of cells, these are the first recordings of the real-time dynamic of glucose movements in a biological microenvironment.     

Improved electrochemical microsensor for the real-time simultaneous analysis of endogenous nitric oxide and carbon monoxide generation

An amperometric dual NO/CO microsensor was developed on the basis of a working electrode incorporating dual Pt microdisks (each diameter, 76 μm) and a Ag/AgCl reference electrode covered with a gas permeable membrane. One of the Pt disks was sequentially electrodeposited with Pt and Sn; the other Pt disk was deposited with Pt-Fe(III) oxide nanocomposites. The first showed activity for the oxidation of both NO and CO; the second showed activity only for NO oxidation. In the copresence of NO and CO, the currents measured at each electrode, respectively, represented the concentrations of CO and NO. The sensor showed high stability during the monitoring of organ tissue for at least 2.5 h and high selectivity to NO over CO at the Pt-Fe(III) oxide working electrode. Real-time coupled dynamic changes of NO and CO generated by a living C57 mouse kidney were monitored simultaneously and quantitatively in response to a NO synthase inhibitor (N(G)-nitro-l-arginine methyl ester), for the first time. CO was found to increase and NO decreased upon addition of the inhibitor, suggesting a possible reciprocal interaction between these endogenous gases.     

A carbon nanotube needle biosensor

A carbon nanotube needle biosensor was developed to provide fast, cost effective and highly sensitive electrochemical detection of biomolecules. The sensor was fabricated based on an array of aligned multi-wall carbon nanotubes synthesized by chemical vapor deposition. A bundle of nanotubes in the array was welded onto the tip of a tungsten needle under a microscope. The needle was then encased in glass and a polymer coating leaving only the tip of the needle exposed. Cyclic voltammetry was performed to examine the redox behavior of the nanotube needle. The cyclic voltammetry results showed a steady-state response attributable to radial diffusion with a high steady-state current density. An amperometric sensor was then developed for glucose detection by physically attaching glucose oxidase on the nanotube needle. The amperometric response of these nanotube needles showed a high sensitivity with a low detection limit. It is expected that the nanotube needle can be sharpened to increase the sensitivity to the point where the current is almost too small to measure. The simple manufacturing method should allow commodity level production of highly sensitive electronic biosensors.     

Manipulation of microenvironment with a built-in electrochemical actuator in proximity of a dissolved oxygen microsensor

Biochemical sensors for continuous monitoring require dependable periodic self diagnosis with acceptable simplicity to check its functionality during operation. An in-situ self-diagnostic technique for a dissolved oxygen microsensor is proposed in an effort to devise an intelligent microsensor system with an integrated electrochemical actuation electrode. With a built-in platinum microelectrode that surrounds the microsensor, two kinds of microenvironments, called the oxygen-saturated or oxygen-depleted phases, can be created by water electrolysis, depending on the polarity. The functionality of the microsensor can be checked during these microenvironment phases. The polarographic oxygen microsensor is fabricated on a flexible polyimide substrate (Kapton) and the feasibility of the proposed concept is demonstrated in a physiological solution. The sensor responds properly during the oxygen-generating and oxygen-depleting phases. The use of these microenvironments for in-situ self-calibration is discussed to achieve functional integration, as well as structural integration, of the microsensor system.     

微传感器最新发展

近年来，微传感器受到国际传感技术界的广泛关注，本文介绍十多个微传感器，包括三轴加速度计，单，双轴加速度计片，表面微机械陀螺（角速度传感器），微惯性导航系统，微磁通门传感器，磁阻传感器，纳米皮拉尼压力传感器，微科氏质量流量计，毫米波图像传感器，GPS手表（1cm^3)，二氧化碳传感器和微/超微角位移传感器，文事简要介绍它们的基本结构。敏感机理，特点等，从中可以看出微传感器已成为传感技术中有重要应用前景的组成部分。     

Simulations of the frequency response of implantable glucose sensors

The response of enzyme electrode glucose sensors implanted in tissues to physiologic blood glucose oscillations is simulated. Models describe both oxygen-based and peroxide-based glucose sensors in spatially homogeneous medium simulating some mass transfer properties of tissue. Pass-through ratios and delays are reported as a function of frequency for the oxygen-based sensor, and the effects on continuous blood glucose monitoring are illustrated using data from the literature. Certain peroxide-based sensor designs may produce common signals for different glucose concentrations, a characteristic not found in oxygen-based sensors. The dynamic response depends on the frequency of glucose oscillation and is sensitive to sensor type, enzyme activity, and diffusional resistance.     

A wireless, remote query glucose biosensor based on a pH-sensitive polymer

This paper describes a wireless, remote query glucose biosensor using a ribbonlike, mass-sensitive magnetoelastic sensor as the transducer. The glucose biosensor is fabricated by first coating the magnetoelastic sensor with a pH-sensitive polymer and upon it a layer of glucose oxidase (GOx). The pH-responsive polymer swells or shrinks, thereby changing mass, respectively, in response to increasing or decreasing pH values. The GOx-catalyzed oxidation of glucose produces gluconic acid, inducing the pH-responsive polymer to shrink, which in turn decreases the polymer mass. In response to a time-varying magnetic field, a magnetoelastic sensor mechanically vibrates at a characteristic resonance frequency, the value of which inversely depends on sensor mass loading. As the magnetoelastic films are magnetostrictive, the vibrations launch magnetic flux that can be remotely detected using a pickup coil. Hence, changes in the resonance frequency of a passive magnetoelastic transducer are detected on a remote query basis, without the use of physical connections to the sensors.The sensitivity of the glucose biosensors decreases with increasing ionic strength; at physiological salt concentrations, 0.6 mmol/L of glucose can be measured. At glucose concentrations of 1-10 mmol/L, the biosensor response is reversible and linear, with the detection limit of 0.6 mmol/L corresponding to an error in resonance frequency determination of 20 Hz. Since no physical connections between the sensor and the monitoring instrument are required, this sensor can potentially be applied to in vivo and in situ measurement of glucose concentrations.     

Modeling the response function of dual-enzyme microbiosensors

A general theoretical model for competitive dual-enzyme microbiosensors based on self-assembled monolayers (SAM) is presented. The model is derived for amperometric dual-enzyme ATP sensors and provides excellent agreement with experimental ATP measurements at 25 microm diameter microelectrodes. In this model, the statistical probability of a glucose molecule in competition between two enzymes, glucose oxidase (GOD)/hexokinase (HEX), at the ATP sensor surface is combined with the enzymatic reaction rate. Thereby, a simple model predicting the sensor signal for varying surface concentrations of GOD and HEX, glucose concentration, and ATP concentration is obtained. Excellent agreement of the predicted current signal with experimentally obtained sensor signals was achieved at ATP concentrations between 10 and 300 microM in a buffer containing glucose at physiologically relevant levels. Consequently, the development time for new dual-enzyme biosensors can be reduced, and an analytical model for the sensor response function is provided facilitating the calibration of enzymatic biosensors.     

Fully Packaged Nonenzymatic Glucose Microsensors With Nanoporous Platinum Electrodes for Anti-Fouling

CMOS integrable nonenzymatic glucose microsensors with nanoporous platinum (Pt) working and counter electrode (WE/CE) were first fabricated, packaged, and characterized with biocompatible Nafion and hydrophilic polyurethane (HPU) membrane materials. Optimal packaging material and its processing condition for these nonenzymatic sensors were investigated. The optimally packaged glucose microsensor was evaluated in human blood plasma solution for checking its biocompatibility and commercial applicability. The fabricated microsensors with nanoporous Pt WE/CE had a sensitivity of 7.75 $mu$A mM $^{-1}$ cm $^{- 2}$. The packaged microsensor with Nafion membrane had better performance characteristics than packaged one with HPU. The packaged microsensor with 1:6 ratio of Nafion to ethanol exhibited a sensitivity of 0.83 $mu$ A mM$^{- 1}$ cm$^{-2}$ and stable current change in the human blood plasma solution, while the current response of nonpackaged microsensor was rapidly saturated because adsorption of various proteins and cells as expected. These data indicate that the packaged nonenzymatic microsensor with biocompatible Nafion membrane is promising and strongly applicable for in vitro and in vivo glucose monitoring systems.      

Information coding in artificial olfaction multisensor arrays

High-density sensor arrays were prepared with microbead vapor sensors to explore and compare the information coded in sensor response profiles following odor stimulus. The coded information in the sensor-odor response profiles, which is used for odor discrimination purposes, was extracted from the microsensor arrays via two different approaches. In the first approach, the responses from individual microsensors were separated (decoded array) and independently processed. In the second approach, response profiles from all microsensors within the entire array, i.e., the sensor ensemble, were combined to create one response per odor stimulus (nondecoded array). Although the amount of response data is markedly reduced in the second approach, the system shows comparable odor discrimination rates for the two signal extraction methods. The ensemble approach streamlines system resources without decreasing system performance. These signal compression approaches may simulate or parallel information coding in the mammalian olfactory system.     

Measurement of tear glucose levels with amperometric glucose biosensor/capillary tube configuration

An amperometric needle-type electrochemical glucose sensor intended for tear glucose measurements is described and employed in conjunction with a 0.84 mm i.d. capillary tube to collect microliter volumes of tear fluid. The sensor is based on immobilizing glucose oxidase on a 0.25 mm o.d. platinum/iridium (Pt/Ir) wire and anodically detecting the liberated hydrogen peroxide from the enzymatic reaction. Inner layers of Nafion and an electropolymerized film of 1,3-diaminobenzene/resorcinol greatly enhance the selectivity for glucose over potential interferences in tear fluid, including ascorbic acid and uric acid. Further, the new sensor is optimized to achieve very low detection limits of 1.5 ± 0.4 μM of glucose (S/N = 3) that is required to monitor glucose levels in tear fluid with a glucose sensitivity of 0.032 ± 0.02 nA/μM (n = 6). Only 4-5 μL of tear fluid in the capillary tube is required when the needle sensor is inserted into the capillary. The glucose sensor was employed to measure tear glucose levels in anesthetized rabbits over an 8 h period while also measuring the blood glucose values. A strong correlation between tear and blood glucose levels was found, suggesting that measurement of tear glucose is a potential noninvasive substitute for blood glucose measurements, and the new sensor configuration could aid in conducting further research in this direction.     

Tuning sensitivity and selectivity of complementary metal oxide semiconductor-based capacitive chemical microsensors

New details on selectivity and sensitivity of fully integrated CMOS-based capacitive chemical microsensor systems are revealed. These microsystems have been developed to detect volatile organics in ambient air and rely on polymeric sensitive layers. The sensitivity and selectivity changes induced by thickness variation of the sensitive polymer layer allow for tuning of the layer parameters to achieve desired sensor features. Cross-sensitivity to interfering agents can be drastically reduced, as is shown for two important cases: (a). rendering the capacitive sensor insensitive to a low-dielectric-constant analyte (lower than that of the polymer) and (b). reducing the influence of a high-dielectric-constant analyte, such as water, on the sensor response. The second case is of vital importance for capacitive sensors, since water is omnipresent and evokes large capacitive sensor signals. The thickness-induced selectivity is explained as a combination of dielectric constant change and swelling and has been confirmed by measurements. Experimentally determined sensitivities qualitatively and quantitatively coincide with the calculated values implying understanding of the sensing mechanism.     

Dual fluorescence sensor for trace oxygen and temperature with unmatched range and sensitivity

An optical dual sensor for oxygen and temperature is presented that is highly oxygen sensitive and covers a broad temperature range. Dual sensing is based on luminescence lifetime measurements. The novel sensor contains two luminescent compounds incorporated into polymer films. The temperature-sensitive dye (ruthenium tris-1,10-phenanthroline) has a highly temperature-dependent luminescence and is incorporated in poly(acrylonitrile) to avoid cross-sensitivity to oxygen. Fullerene C70 was used as the oxygen-sensitive probe owing to its strong thermally activated delayed fluorescence at elevated temperatures that is extremely oxygen sensitive. The cross-sensitivity of C70 to temperature is accounted for by means of the temperature sensor. C70 is incorporated into a highly oxygen-permeable polymer, either ethyl cellulose or organosilica. The two luminescent probes have different emission spectra and decay times, and their emissions can be discriminated using both parameters. Spatially resolved sensing is achieved by means of fluorescence lifetime imaging. The response times of the sensor to oxygen are short. The dual sensor exhibits a temperature operation range between at least 0 and 120 degrees C, and detection limits for oxygen in the ppbv range, operating for oxygen concentrations up to at least 50 ppmv. These ranges outperform all dual oxygen and temperature sensors reported so far. The dual sensor presented in this study is especially appropriate for measurements under extreme conditions such as high temperatures and ultralow oxygen levels. This dual sensor is a key step forward in a number of scientifically or commercially important applications including food packaging, for monitoring of hyperthermophilic microorganisms, in space technology, and safety and security applications in terms of detection of oxygen leaks.     

Oxygen microsensor and its application to single cells and mouse pancreatic islets.

An oxygen microsensor with a < 3-micron tip diameter was developed for monitoring oxygen levels at single cells and mouse pancreatic islets. The sensor was fabricated by electrochemically recessing an etched Pt wire inside a pulled glass micropipet and then coating with cellulose acetate. This fabrication process was found to be simpler than previous oxygen electrode designs of comparable size. The microsensors had a average sensitivity of 0.59 +/- 0.29 pA/mmHg (mean +/- SD, n = 42), signals that were minimally perturbed by convection, and response times of < 1 s. The electrode was used to measure the oxygen gradient around and inside single mouse islets. The measurements demonstrate that oxygen levels within even the largest islets at maximal glucose stimulation are 67 +/- 1.6 mmHg (mean +/- SD, n = 5), indicating that islets have adequate oxygen supplies by diffusion under tissue culture conditions to support insulin secretion. The electrode was also used to record the dynamics of oxygen level at single islets as a function of glucose concentration. As glucose level was changed from 3 to 10 mM, oxygen level decreased by 15.8 +/- 2.3 mmHg (mean +/- SEM, n = 6) and oscillations with a period of 3.3 +/- 0.6 min (mean +/- SEM, n = 6) appeared in the oxygen level. In islets bathed in quiescent solutions containing 10 mM glucose, similar oscillations could be observed. In addition, in the quiet solutions it was possible to detect faster oscillations with a period of 12.1 +/- 1.7 s (mean +/- SEM, n = 6) superimposed on the slower oscillations. Oxygen consumption could also be observed at single insulinoma cells using the electrode. Individual cells also showed oscillations in oxygen consumption with a period of a few seconds. The results demonstrate that the electrode can be used for dynamic oxygen level recordings in biological microenvironments.     

Glucose-sensitive holographic sensors for monitoring bacterial growth

A glucose sensor comprising a reflection hologram incorporated into a thin, acrylamide hydrogel film bearing the cis-diol binding ligand, 3-acrylamidophenylboronic acid (3-APB), is described. The diffraction wavelength (color) of the hologram changes as the polymer swells upon binding cis-diols. The effect of various concentrations of glucose, a variety of mono- and disaccharides, and the alpha-hydroxy acid, lactate, on the holographic response was investigated. The sensor displayed reversible changes in diffraction wavelength as a function of cis-diol concentration, with the sensitivity of the system being dependent on the cis-diol tested. The effect of varying 3-APB concentration in the hydrogel on the holographic response to glucose was investigated, and maximum sensitivity was observed at a functional monomer concentration of 20 mol %. The potential for using this holographic sensor to detect real-time changes in bacterial cell metabolism was demonstrated by monitoring the germination and subsequent vegetative growth of Bacillus subtilis spores.     

用于氨氮检测的无透气膜安培型氨气微传感器及系统

设计并制备了一种无透气膜的安培型氨气微传感器,并构建了氨氮检测系统,该系统实现了将溶液中的氨氮转化为氨气,并应用所设计的微传感器检测气态氨氮．采用MEMS工艺制备组成该微传感器的微电极芯片,氨敏感材料选用铂黑,通过电化学方法修饰于微电极表面．在微电极芯片的SU-8微池中滴入微量LiCl溶液,形成可使氨气迅速扩散到电极表面的薄层电解液．使用自行设计的氨氮检测系统对不同浓度氨氮样品进行检测,对微传感器的时间响应特性、线性度、灵敏度及重复性进行了测试和分析．微传感器的线性范围为0．2mg／L～4mg／L,线性相关系数为0．9847,检测下限为0．2mg／L,达到90％响应信号所需的时间在3min以内．结果表明,使用安培型氨气微传感器检测氨氮的方法是可行的．