Flow-Cell Fibre-Optic Enzyme Sensor for Phenols

Abstract

A solid-state fibre-optic luminescent oxygen sensor was used for flow-through measurements. It acts as a transducer in a new flow-cell enzyme sensor arrangement. This arrangement comprises a flow path, sample injector, microcolumn with the immobilized enzyme, oxygen membrane and fibre-optic connector joined together to form an integral unit. Laccase enzyme was used as a recognition system which provided specific oxidation of the substrates with the dissolved oxygen being monitored. The assay procedure was optimized and performance of the new system studied. The sensor was applied to the determination polyphenol content in tea, brandy, etc. (quality control test). The sensitivity to some important phenolic compounds was tested with the view of industrial wastewater control applications.     

High sensitivity in gas analysis with photoacoustic detection

Introduction of a new type of pressure sensor has been shown to improve orders of magnitude the sensitivity of a photoacoustic measurement system using a black body radiation source. A new pressure sensor was developed to overcome the limitations in the capacitive microphone technology and to obtain ultimate sensitivity in photoacoustic gas detection when using low modulation frequency below 500 Hz. The pressure sensor is a cantilever-type microphone with interferometric measurement of the sensor displacement. By using conventional filter-type photoacoustic setup with the cantilever microphone and a black body radiation source, we obtained a detection limit in the sub-ppb range for methane gas with 100 s measurement time.     

A Surface Plasmon Resonance Sensor Platform Coupled with Gold Nanoparticle Probes for Unpurified Nucleic Acids Detection

In this work, a surface plasmon resonance sensor system was designed and implemented for determination of nucleic acids in unpurified samples. First, through blocking non-specific interaction sites on the sensor surface to reduce non-specific adsorption from unpurified sample matrix, it was determined that at the optimal BSA concentration of 100 µg/ml the non-specific interaction can be reduced by 50%, although improvement for direct detection of nucleic acids in unpurified sample is required. Second, bearing nonspecific adsorption onto gold films, nucleic acids adsorbed on sensor surface in unpurified sample matrix were detected through a secondary hybridization approach. Using DNA-lined AuNPs shows the new SPR sensor can be applied for the determination of target ssDNA with a detection range of 0.1–10 µM for targets in purified and 1–10 µM in unpurified samples, respectively. Results imply that the new SPR sensor system is promising for specific and convenient analysis of nucleic acids directly in unpurified samples. Development of the new SPR sensor technique can have applications in fast field diagnostics and monitoring.     

Solid state biosensors using thin-film electrodes

Solid-state technology was used to fabricate a new type of glucose sensor and a differential measurement system was incorporated with the sensor in order to suppress interfering signals. This glucose sensor was composed of film electrodes and an enzyme-immobilizing membrane. The hydrogen peroxide electrodes were fabricated by metallic film deposition. Glucose oxidase (GOD), an enzyme, was immobilized in photosensitive polyvinyl alcohol to enable the enzyme-immobilizing membrane to be formed by the usual photolithographic method. Two anodes and one common cathode were formed on a glass substrate. One of the anodes was coated with PVA containing active enzyme and the other with PVA containing deactivated enzyme. By setting up a differential output measurement system, it was possible to suppress the interfering signals that might be generated by some other reducing substances in a sample solution. With this measuring technique, the signal of ascorbic or uric acids up to 100 mg/dl was successfully cancelled. Glucose concentrations in the range 5 to 100 mg/dl could be detected by the present sensor.The fabrication processes of the present sensor were so compatible with IC technology that it should be possible to mass produce disposable glucose biosensors using IC fabrication techniques.     

基于多壁碳纳米管/纳米铜复合材料的电化学传感器测定槐米中芦丁

以Nafion-多壁碳纳米管(MWNTs)薄膜修饰玻碳电极,采用恒电位法在其表面电沉积纳米Cu,研制了一种新型的芦丁电化学传感器。采用循环伏安法和电化学阻抗谱研究传感器在铁氰化钾-亚铁氰化钾体系中的电化学行为,以考察传感器的电化学性能,采用扫描电镜对传感器表面的形态进行了研究,并利用差分脉冲伏安法测定芦丁的含量。实验结果表明,该传感器对芦丁有较好的催化作用。在优化实验条件下,对芦丁检测的线性范围为1.0×10-8~1.0×10-6 mol/L,检出限为8.4×10-9 mol/L(S/N=3),回收率为98.1%~101%,该传感器制作简单,线性范围宽,灵敏度高,为槐米等样品中芦丁含量的检测提供了一种新的、行之有效的方法。     

基于电化学电容检测的新型钙离子传感器

通过金-硫共价键,将巯基十六羧酸以自组装的方式固定到金电极表面。再通过EDC、NHS的活化作用,对钙调蛋白的氨基与巯基十六羧酸暴露于自组装层外的羧基进行共价键和,从而将钙调蛋白固定到电极表面,制成一种新型的钙离子电容传感器。首先对传感器进行了等效电路分析,并分别考察了不同浓度电解质溶液对基底电容的影响,确定了进行实验所需的合适电解质浓度。用本实验室自行开发研制的电化学电容分析仪研究了该传感器对钙离子的响应能力,并进一步考察了其再生能力及稳定性。实验结果表明,在10-8~3×10-6mol/L的较宽范围内,钙离子浓度的对数值与总电容信号值之间呈线性关系,而且检出限可以达到10-9mol/L。制备好的传感器浸泡在0.02 mol/L,pH 7.0的磷酸缓冲溶液中在4℃冰箱中可以保存一个月而其对钙离子的响应能力未发现有显著的下降,说明其具有良好的稳定性。使用过的传感器用EDTA进行再生后基底电容的变化不超过0.3%,说明其具有良好的再生可逆性,可以被重复利用。本工作为对钙离子进行定量检测提供了一种新方法。     

Organelle-specific zinc detection using zinpyr-labeled fusion proteins in live cells

A protein labeling approach is employed for the localization of a zinc-responsive fluorescent probe in the mitochondria and in the Golgi apparatus of living cells. ZP1, a zinc sensor of the Zinpyr family, was functionalized with a benzylguanine moiety and thus converted into a substrate (ZP1BG) for the human DNA repair enzyme alkylguaninetransferase (AGT or SNAP-Tag). The labeling reaction of purified glutathione S-transferase tagged AGT with ZP1BG and the zinc response of the resulting protein-bound sensor were confirmed in vitro. The new detection system, which combines a protein labeling methodology with a zinc fluorescent sensor, was tested in live HeLa cells expressing AGT in specific locations. The enzyme was genetically fused to site-directing proteins that anchor the probe onto targeted organelles. Localization of the zinc sensors in the Golgi apparatus and in the mitochondria was demonstrated by fluorescence microscopy. The protein-bound fluorescence detection system is zinc-responsive in living cells.     

平面波导型荧光免疫传感器的制备与应用

研究了一种基于全内反射荧光和免疫检测原理的传感器系统,它可用于检测水环境中的有机污染物,如2,4-D、藻毒素等.本系统采用长波长的荧光染料Cy 5.5,以氦氖激光器作为激发光源,结合流动注射分析系统,能够快速方便地对传感基片上发生的免疫反应进行测定.本研究介绍了平面波导型荧光免疫传感器的检测原理、系统结构设计、传感基片的修饰、小分子配基的固定和系统测试结果.实验结果表明,系统具有很高的稳定性,对荧光染料的响应灵敏度可达1nmol/L;修饰后的传感基片基本不对蛋白产生非特异性吸附,而对不同浓度小分子配基抗体的响应符合Logistic方程;传感基片重复使用20个周期后,性能没有明显的下降.     

A study of a new TSM bio-mimetic sensor using a molecularly imprinted polymer coating and its application for the determination of nicotine in human serum and urine

A new bio-mimetic quartz crystal thickness-shear-mode (TSM) sensor, using an imprinted polymer coating as the sensitive material, has been fabricated and applied to the determination of nicotine (NIC) in human serum and urine. The molecularly imprinted polymer (MIP) was synthesized using NIC as the template molecule and methacrylic acid (MAA) as the functional monomer. The sensor showed high selectivity and a sensitive response to NIC in aqueous system. The linear response range of the sensor was between 5.0 x 10(-8) and 1.0 x 10(-4) M with a detection limit of 2.5 x 10(-8) M. The viscoelasticity of the coating in the air and in liquid has been studied by the impedance spectrum. The MIP sensor was stable and exhibited effective reproducibility. Satisfactory results were achieved in the detection of the real samples.     

Development of a modified grating coupler in application to geosciences

A grating coupler system has been developed to measure refractive index gradients with high spatial (6.7 μm) and temporal (milliseconds) resolution. The system was applied to two-phase model systems consisting of water and non-aqueous pollution liquids. Refractive index gradients at the interfaces between the aqueous and organic phase of 1-butanol, hexane, and 1-heptanol were monitored under steady-state conditions. The temporal resolution was utilized in diffusion experiments with glycerol and sodium chloride in water, where the formation of a concentration gradient was studied. In a further application, the grating coupler system was modified to monitor low-level concentrations of aqueous pollution profiles as are caused by bacterial degradation in the aqueous phase. Toluene was selected as contaminant. The sensor sensitivity was improved by coating the sensor with the pre-concentrating polymers polydimethylsiloxane and Teflon® AF-2400. With the grating coupler setup, a multi-purpose instrument was created to measure high-resolution refractive index gradients with high temporal and spatial resolution in different fields of application. The new sensor system can be used to measure absolute refractive indices by covering parts of the sensing area with cover media of known refractive index. Coatings can be used for sensitivity improvement by pre-concentrating the sample, for selectivity by utilizing filtering properties of the coating, and as calibration standard for absolute refractive index measurements.     

Glutaric Acid Assisted Fabrication of CuO Nanostructures and their Application in Development of Highly Sensitive Electrochemical Sensor System for Carbamates

This work describes the fabrication of unique arrow head shaped CuO nanostructures using simple hydrothermal treatment method. The highly attractive features were obtained by the application of glutaric acid utilised simultaneous as template and functionalising agent. The functionalised nanostructures were known to possess excellent potential towards the electro‐catalytic oxidation of carbofuran pesticide. The generated intense electrochemical signal with lower potential value enabled sensitive and selective determination of carbofuran up to 1×10^?3 µM with wide sensing window in range of 0.01 to 0.16 µM. The feasibility of the developed sensor system for the practical application was also studied by testing its potential in real sample extracts of various vegetables. The excellent recoveries demonstrated the analytical robustness of the developed sensor system. The sensor system utilises a new and simple approach towards sensitive determination of toxic pesticides reflecting its wide spectrum application in various fields.     

An Electrochemiluminesence Chiral Sensor for Propranolol Enantiomers Based on Functionalized Graphite‐like Carbon Nitride Nanosheets

Graphite‐like carbon nitride nanosheets (g‐C₃N₄) have been attracted considerable attention for their applications in catalysis and electrochemiluminesence (ECL) sensor. In this paper, a facile solvothermal method was employed to prepare the functionalized nanocomposites with metal salts cadmium carbonate (CdCO₃) and g‐C₃N₄ hybrids (g‐C₃N₄‐CdCO₃). The prepared materials were characterized by scanning electron microscope (SEM), X‐ray energy dispersive spectroscopy (EDX) and Fourier transform infrared spectra (FTIR). The nanocomposites was used not only as a new type luminophore but also as a chiral selector in this simple and sensitive ECL chiral sensor system for the recognition and detection of propranolol (Pro) enantiomers via quenching effect. The obvious difference of ECL signal of S‐Pro and R‐Pro was obtained. Thus, the sensor had the ability to distinguish Pro enantiomers. The mechanism of recognition was discussed by the theoretical calculation of binding constant and the water contact angle experiments. The sensor for Pro enantiomers was developed at wide concentration range from 0.001 to 1 mmol?L^?1. The sensor with excellent sensitivity, stability and reproducibility provide a new strategy based on ECL for chiral recognition and the ECL chiral sensor.     

Fiber-optic flow-through sensor for online monitoring of glucose

A new microdialysis-based glucose-sensing system with an integrated fiber-optic hybrid sensor is presented. Design and dimensions of the cell are adapted for its coupling with commercially available microdialysis techniques, thereby providing a new system for continuous glucose monitoring. The glucose level is detected via oxygen consumption which occurs as a consequence of enzymatic reaction between immobilized glucose oxidase and glucose. The use of gas-permeable Tygon tubing ensures complete and constant air-saturation of the measuring fluid in the cell. Nevertheless, a reference oxygen optode is used to detect and to compensate response changes caused by events like bacterial growth, temperature fluctuations, or failure of the peristaltic pump. In contrast to widely used electrochemical sensors, the response of the microdialysis-based fiber-optic glucose sensor is highly selective, making this sensor approach particularly advantageous for continuous glucose monitoring of patients in intensive care units. The effects of flow rate, pH, temperature, and common interferences on the sensor response are presented and discussed in detail. The sensor is evaluated in vitro using a 3-day continuous test in glucose-spiked plasma. The ability to measure glucose in humans is demonstrated by coupling the flow-through cell and commercially available microdialysis catheter CMA60. A 24-h monitoring test using this setup is successfully applied to a healthy volunteer.     

Modified Natural Rubber as a Simple Chemical Sensor with Smartphone Detection for Formaldehyde Content in a Seafood Sample

A new biodegradable platform-based sensor for formaldehyde assay is proposed. Natural rubber latex was modified to polylactic acid–chloroacetated natural rubber polymer blend sheets. The polymer blend sheet was grafted using a water-based system with amine monomers as a platform, with a spot exhibiting positive polarity for immobilizing with anionic dye (Acid Red 27). The sensor was exposed to formaldehyde. The color intensity of the dye on the sensor spot would decrease. Using a smartphone with image processing (via ImageJ program), the color intensity change (∆B) could be followed. A linear calibration, ∆B intensity = 0.365 [FA] + 6.988, R² = 0.997, was obtained for 10–150 mM FA with LOD and LOQ at 3 and 10 mM, respectively (linear regression method). The precision was lower than 20% RSD. Application to real seafood samples was demonstrated. The ready-to-use sensor with the proposed method was cost-effective, was portable for on-site analysis, and demonstrated green chemical analysis.     

一种微机控制的自动光度滴定系统

本文介绍了作者研制光度滴定传感器，它具有较小的工作电流，较高的灵敏度和较低的成本。     

A surface plasmon resonance immunosensor for detecting a dioxin precursor using a gold binding polypeptide

A surface plasmon resonance (SPR) based biosensor was developed for monitoring 2,4-dichlorophenol, a known dioxin precursor, using an indirect competitive immunoassay. The SPR sensor was fabricated by immobilizing a gold-thin layer on the surface of an SPR sensor chip with an anti-(2,4-dichlorophenol) antibody using a gold binding polypeptide (GBP) and protein G. The SPR response based on the antigen-antibody reaction in a flow system was measured by injecting a 2,4-dichlorophenol sample solution into the flow system in which the SPR sensor was located. In a direct immunoassay system using the modified sensor chip, no significant SPR angle shift less than 0.001° was observed when a 25 ppm of 2,4-dichlorophenol solution was injected. In order to improve the sensitivity of the SPR sensor, an indirect competitive immunoassay method was used in conjunction with the SPR sensor system using 2,4-dichlorophenol conjugated with bovine serum albumin (BSA). In the competitive assay, a 350 ppm 2,4-dichlorophenol-BSA conjugate solution containing 2,4-dichlorophenol at various concentrations (10-250 ppb) were injected into the SPR sensor system. The sensitivity of this indirect immunoassay was found to be extremely sensitive, compared to the direct one, and a detection limit of 20 ppb was estimated. Verification that the use of GBP for immobilizing the antibody on the sensor chip enhanced the sensitivity to 2,4-dichlorophenol was obtained by comparing the procedure with another modification, in which BSA was used instead of GBP for immobilizing the antibody on the sensor chip. The affinity constant of 2,4-dichlorophenol and its conjugate to the antibody were estimated form the SPR response.     

Oligonucleotide-functionalized gold nanoparticles-enhanced QCM-D sensor for mercury(II) ions with high sensitivity and tunable dynamic range

A quartz crystal microbalance with dissipation monitoring (QCM-D) sensor was developed for highly sensitive and specific detection of mercury(II) ions (Hg(2+)) with a tunable dynamic range, using oligonucleotide-functionalized gold nanoparticles (GNPs) for both frequency and dissipation amplification. The fabrication of the sensor employed a 'sandwich-type' strategy, and formation of T-Hg(2+)-T structures in linker DNA reduced the hybridization of the GNPs-tagged DNA on the gold electrode, which could be used as the molecular switch for Hg(2+) sensing. This QCM-D mercury sensor showed a linear response of 10-200 nM, with detection limits of 4 nM and 7 nM for frequency and dissipation measurements, respectively. Moreover, the dynamic range of the sensor could be tuned by simply altering the concentration of linker DNA without designing new sensors in the cases where detection of Hg(2+) at different levels is required. This sensor afforded excellent selectivity toward Hg(2+) compared with other potential coexisting metal ions. The feasibility of the sensor was demonstrated by analyzing Hg(2+)-spiked tap- and lake-water samples with satisfactory recoveries. The proposed approach extended the application of the QCM-D system in metal ions sensing, and could be adopted for the detection of other analytes when complemented with the use of functional DNA structures.     

Cetyltrimethylammonium Bromide(CTAB)-Ionic Liquid Composite Modified Electrode for Sensitive Cyclic Voltammetric Determination of Bisphenol A

A sensitive electrochemical sensor for determining bisphenol A(BPA) was designed. The sensor was a glassy carbon electrode modified with the surfactant cetyltrimethylammonium bromide and the ionic liquid 1-decyl-3-methylimidazolium tetrafluoroborate. The ability of the new sensor to measure BPA was investigated in cyclic voltammetry experiments. Under optimized conditions, the sensor gave a linear response range for BPA of 2.19×10^-7-3.28×10^-5 mol/L and a detection limit of 7.31×10^-8 mol/L(S/N=3). BPA could be determined with a lower detection limit, a wider linear range, and more sensitivity using the sensor than using other electrochemical sensors or high performance liquid chromatography with UV detection. The new sensor was used to determine BPA in tap water with recoveries of 97.5%-98.7% and a relative standard deviation <2.9%. The results show that the sensor can be used to determine trace BPA concentrations in tap water.     

Colorimetric detection of sodium ion in serum based on the G-quadruplex conformation related DNAzyme activity

There has been a big challenge in developing the Na⁺ sensor that can be practically used in the physiological system with the interference of large amounts of K⁺. In this research, a novel Na⁺ sensor has been designed based on the G-quadruplex-conformation related DNAzyme activity. The sensor exhibits high selectivity and sensitivity with the detection limit of 0.6 μM, which enables the sensor to be practically used in determination of the Na⁺ level in serum. The research not only provides a simple Na⁺ sensor but also opens a new way for developing the detection technology of Na⁺.     

Fluorescence monitoring of a benzo[a]pyrene metabolite using a regenerable immunochemical-based fiber-optic sensor

Microscale regenerable biosensors are described and utilized to measure the natural fluorophor benzo[a]pyrene tetraol (BPT). The sensors combine laser-excited/fiber-optic remote sensing principles with a unique capillary tube delivery system to make repetitive, heterogeneous fluoroimmunoassay measurements. Two sensor configurations and modes of operation are described. Concentrations of BPT in the nanomolar range are easily measured with a reproducibility of 10% or better, depending on the sensor design, selective measurements can be made in ca. 20 min, then the sensor can be regenerated by delivering new reagents to the sensing chamber, without removing the sensor from the sample.