Fully enclosed microfluidic paper-based analytical devices

This article introduces fully enclosed microfluidic paper-based analytical devices (microPADs) fabricated by printing toner on the top and bottom of the devices using a laser printer. Enclosing paper-based microfluidic channels protects the channels from contamination, contains and protects reagents stored on the device, contains fluids within the channels so that microPADs can be handled and operated more easily, and reduces evaporation of solutions from the channels. These benefits extend the capabilities of microPADs for applications as low-cost point-of-care diagnostic devices.     

Applications of polymers for biomolecule immobilization in electrochemical biosensors

Polymers are becoming inseparable from biomolecule immobilization strategies and biosensor platforms. Their original role as electrical insulators has been progressively substituted by their electrical conductive abilities, which opens a new and broad scope of applications. In addition, recent advances in diagnostic chips and microfluidic systems, together with the requirements of mass-production technologies, have raised the need to replace glass by polymeric materials, which are more suitable for production through simple manufacturing processes. Conducting polymers (CPs), in particular, are especially amenable for electrochemical biosensor development for providing biomolecule immobilization and for rapid electron transfer. It is expected that the combination of known polymer substrates, but also new transducing and biocompatible interfaces, with nanobiotechnological structures, like nanoparticles, carbon nanotubes (CNTs) and nanoengineered ‘smart’ polymers, may generate composites with new and interesting properties, providing higher sensitivity and stability of the immobilized molecules, thus constituting the basis for new and improved analytical devices for biomedical and other applications. This review covers the state-of-the-art and main novelties about the use of polymers for immobilization of biomolecules in electrochemical biosensor platforms.     

On-chip surface-based detection with nanohole arrays

A microfluidic device with integrated surface plasmon resonance (SPR) chemical and biological sensors based on arrays of nanoholes in gold films is demonstrated. Widespread use of SPR for surface analysis in laboratories has not translated to microfluidic analytical chip platforms, in part due to challenges associated with scaling down the optics and the surface area required for common reflection mode operation. The resonant enhancement of light transmission through subwavelength apertures in a metallic film suggests the use of nanohole arrays as miniaturized SPR-based sensing elements. The device presented here takes advantage of the unique properties of nanohole arrays: surface-based sensitivity; transmission mode operation; a relatively small footprint; and repeatability. Proof-of-concept measurements performed on-chip indicated a response to small changes in refractive index at the array surfaces. A sensitivity of 333 nm per refractive index unit was demonstrated with the integrated device. The device was also applied to detect spatial microfluidic concentration gradients and to monitor a biochemical affinity process involving the biotin-streptavidin system. Results indicate the efficacy of nanohole arrays as surface plasmon-based sensing elements in a microfluidic platform, adding unique surface-sensitive diagnostic capabilities to the existing suite of microfluidic-based analytical tools.     

Electrophoretic cell manipulation and electrochemical gene-function analysis based on a yeast two-hybrid system in a microfluidic device

A novel microfluidic device with an array of analytical chambers was developed in order to perform single-cell-based gene-function analysis. A series of analytical processes was carried out using the device, including electrophoretic manipulation of single cells and electrochemical measurement of gene function. A poly(dimethylsiloxane) microstructure with a microfluidic channel (150 microm in width, 10 microm in height) and an analytical chamber (100 x 20 x 10 microm (3)) were fabricated and aligned on a glass substrate with an array of Au microelectrodes. Two microelectrodes positioned in the analytical chamber were employed as a working electrode for the electrophoretic manipulation of cells and electrochemical measurements. A yeast strain ( Saccharomyces cerevisiae Y190) carrying the beta-galactosidase reporter gene was used to demonstrate that the device could detect the enzyme. Target cells flowing through the main channel were introduced into the chamber by electrophoresis using the ground electrode laid on the main channel. When the cell was treated with 17beta-estradiol, gene expression was triggered to produce beta-galactosidase, catalyzing the hydrolysis of p-aminophenyl-beta- D-galactopyranoside to form p-aminophenol (PAP). The enzymatically generated PAP was detected by cyclic voltammetry and amperometry at the single-cell level in the chamber of the device. Generator-collector mode amperometry was also applied to amplify the current response originating from gene expression in the trapped single cells. After electrochemical measurement, the trapped cells were easily released from the chamber using electrophoretic force.     

Electrochemical detection of testosterone by use of three-dimensional disc-ring microelectrode sensing platforms: application to doping monitoring

Testosterone is one of the androgenic steroid hormones, the consumption of which is considered doping in most sports. Here, we present powerful 3D sensing platforms using novel disc-ring microelectrode array devices and exploit them for the competitive immunosensing of testosterone. Each device contains a microelectrode array that consists of a large number of individual microdiscs and is used as the substrate for immunofunctionalization and assay performance. One micrometer above it, a second microelectrode array, this time consisting of microrings, is used as the working electrode for electrochemical monitoring. The physical separation of these two functions allows the incorporation of relatively thick biocomponent layers during immunofunctionalization of the microdiscs without negatively affecting electrochemical detection at the rings. Moreover, it permits electrochemical activation of the latter immediately before substrate addition and hence enables optimal electrode performance. The optimized assay showed a linear range between 0.01 and 10 ng/mL and a limit of detection of 12.5 pg/mL testosterone with detection times of 45 min.     

On-chip electric field driven electrochemical detection using a poly(dimethylsiloxane) microchannel with gold microband electrodes

An external electric field driven in-channel detection technique for on-chip electrochemical detection in micro fabricated devices is described based on a microfluidic system containing an array of 20 microband electrodes. It is shown that an external electric field induces a potential difference between two gold microband electrodes in a poly(dimethylsiloxane) (PDMS) microchannel, and that this enables the electrochemical detection of electroactive species such as ascorbic acid and Fe(CN) 6 (4-). The results, which are supported by simulations of the behavior of the microband electrodes in the microfluidic system, show that the induced potential difference between the electrodes can be controlled by altering the external electric field or by using different microbands in the microband array. As the obtained currents depend on the concentrations of electroactive species in the flowing solution and the detection can be carried out anywhere within the channel without interference of the external electric field, the present approach significantly facilitates electrochemical detection in capillary electrophoresis. This approach consequently holds great promise for application in inexpensive portable chip-based capillary electrophoresis (CE) devices.     

Preparation and electrochemical application of a new biosensor based on plant tissue/polypyrrole for determination of acetaminophen

Banana tissue containing polyphenol oxidase was incorporated into polypyrrole matrix to make a biosensor for the analysis of acetaminophen (ACT). The electrocatalytic behaviour of oxidized acetaminophen was studied at the surface of the biosensor, using various electrochemical methods. The advantages of this biosensor for the determination of acetaminophen are excellent catalytic activity, good detection limit and high exchange current density. The electrochemical and structural properties of the electrode were assessed using cyclic voltammetry, differential voltammetry, chronoamperometric techniques. The analytical properties (sensitivity, I _p ) of this biosensor increased with plant tissue loading. Also this new biosensor was successfully applied for determination of acetaminophen in biologic samples.     

Design of the polyacrylonitrile-reduced graphene oxide nanocomposite-based non-enzymatic electrochemical biosensor for glucose detection

With the advantages of developed electronic devices, various biosensor applications have become attractive issues with excellent electrochemical performances against biomarkers and molecules in biomedical applications. In this study, novel polyacrylonitrile (PAN)-reduced graphene oxide (rGO) nanocomposite-based non-enzymatic electrochemical biosensors were prepared to investigate the detection performance of the glucose. The PAN-rGO nanocomposite-based biosensor detected glucose with a high sensitivity and stability due to enhanced redox mechanism arising from rGO additive. PAN-rGO nanocomposite-based biosensor detected glucose in (0.75–12) mM with a high sensitivity of 49 µAmM^?1 cm^?2 (2.5 times higher than PAN-based sensor). Concentration–response graphs correlating the non-enzymatic electrochemical signal to glucose concentration revealed a low limit of detection (LOD) of 0.6 mM within 1-min voltammetric cycle. The selectivity results confirmed a significant preferential response of the proposed PAN-rGO nanocomposite-based biosensor for glucose against possible interfering compounds. The proposed PAN-rGO nanocomposite-based biosensor has a great potential to be used as a nanostructured platform for detection of glucose in phosphate-buffered saline (pH 7.4) solution with high sensitivity, selectivity, stability, reproducibility, and fast response properties.

     

Stochastic sensing on a modular chip containing a single-ion channel

Engineered protein channels have many potential applications in biosensing at the single-molecule level. A future generation of biosensor could be an array of target-specific ion channels, where each protein pore acts as a sensor element. An important step toward this goal is to create a portable, durable, single-protein channel-integrated chip device. Here we report a versatile, modular chip that contains a single-ion channel for single-molecular biosensing. The core of the device is a long-lived lipid membrane that has been sandwiched between two air-insulated agarose layers which gel in situ. A single-protein pore embedded in the membrane serves as the sensor element. The modular device is highly portable, allowing a single-ion channel to continuously function following detachment of the chip from the instrument and independent transportation of the device. The chip also exhibits high durability, which is evidenced from long-duration continuous observation of single-channel dynamics. Once engineered protein pores are installed, the chip becomes a robust stochastic sensor for real-time targeting such as detection of the second messenger IP3. This pluggable biochip could be incorporated with many applicable devices, such as a microfluidic system, and be made into a microarray for both biomedical detection and membrane protein research.     

Microfluidic separation and gateable fraction collection for mass-limited samples

Integrating multiple analytical processes into microfluidic devices is an important research area required for a variety of microchip-based analyses. A microfluidic system is described that achieves preparative separations by intelligent fraction collection of attomole quantities of sample. The device consists of a main microfluidic channel used to perform electrophoresis, which is interconnected at 90 degrees to two vertically displaced channels via a nanocapillary array membrane. The membrane interconnect contains nanometer-diameter pores that provide fluidic communication between the channels. Sample injection and analyte collection are controlled by application of an electrical bias between the microfluidic channels across the nanocapillary array. After the separation, the automated transfer of the FITC-labeled Arg, Gln, and Gly bands occurs; a fluorescence detector located at the separation/collection channel interconnect is used to generate a triggering signal that initiates suitable voltages to allow near-quantitative transfer of analyte from the separation channel to the second fluidic layer. The ability to achieve such sample manipulations from mass-limited samples enables a variety of postseparation processing events.     

Theory, fabrication and applications of microfluidic and nanofluidic biosensors

Biosensors are a broad array of devices that detect the type and amount of a biological species or biomolecule. Several different types of biosensors have been developed that rely on changes to mechanical, chemical or electrical properties of the transduction or sensing element to induce a measurable signal. Often, a biosensor will integrate several functions or unit operations such as sample extraction, manipulation and detection on a single platform. This review begins with an overview of the current state-of-the-art biosensor field. Next, the review delves into a special class of biosensors that rely on microfluidics and nanofluidics by presenting the underlying theory, fabrication and several examples and applications of microfluidic and nanofluidic sensors.     

Sensitive determination of uric acid by using graphene quantum dots as a new substrate for immobilisation of uric oxidase

A novel strategy for highly sensitive electrochemical detection of uric acid (UA) was proposed based on graphene quantum dots (GQDs), GQDs were introduced as a suitable substrate for enzyme immobilisation. Uric oxidase (UOx) was immobilised on GQDs modified glassy carbon electrode (GCE). Transmission electron microscope, scanning electron microscopy, cyclic voltammetry and electrochemical impedance spectroscopy techniques were used for characterising the electrochemical biosensor. The developed biosensor responds efficiently to UA presence over the concentration linear range 1–800 μM with the detection limit 0.3 μM. This novel biosensing platform based on UOx/GQDs electrode responded even more sensitively than that based on GCE modified by UOx alone. The inexpensive, reliable and sensitive sensing platform based on UOx/GQDs electrode provides wide potential applications in clinical.Inspec keywords: organic compounds, graphene devices, quantum dots, enzymes, biosensors, biochemistry, electrochemical electrodes, electrochemical sensors, transmission electron microscopy, scanning electron microscopy, voltammetry (chemical analysis), electrochemical impedance spectroscopy, nanomedicine, molecular biophysicsOther keywords: sensitive uric acid determination, graphene quantum dots, uric oxidase immobilisation, electrochemical detection, GQD, enzyme immobilisation, glassy carbon electrode, GCE, transmission electron microscope, scanning electron microscopy, cyclic voltammetry, electrochemical impedance spectroscopy, electrochemical biosensor, C     

Highly Sensitive Detection of Protein Biomarkers with Organic Electrochemical Transistors

The analysis of protein biomarkers is of great importance in the diagnosis of diseases. Although many convenient and low‐cost electrochemical approaches have been extensively investigated, they are not sensitive enough in the detection of protein biomarkers with low concentrations in physiological environments. Here, this study reports a novel organic‐electrochemical‐transistor‐based biosensor that can successfully detect cancer protein biomarkers with ultrahigh sensitivity. The devices are operated by detecting electrochemical activity on gate electrodes, which is dependent on the concentrations of proteins labeled with catalytic nanoprobes. The protein sensors can specifically detect a cancer biomarker, human epidermal growth factor receptor 2, down to the concentration of 10^?14 g mL^?1, which is several orders of magnitude lower than the detection limits of previously reported electrochemical approaches. Moreover, the devices can successfully differentiate breast cancer cells from normal cells at various concentrations. The ultrahigh sensitivity of the protein sensors is attributed to the inherent amplification function of the organic electrochemical transistors. This work paves a way for developing highly sensitive and low‐cost biosensors for the detection of various protein biomarkers in clinical analysis in the future.     

A Nanostructured Microfluidic Immunoassay Platform for Highly Sensitive Infectious Pathogen Detection

Rapid and simultaneous detection of multiple potential pathogens by portable devices can facilitate early diagnosis of infectious diseases, and allow for rapid and effective implementation of disease prevention and treatment measures. The development of a ZnO nanorod integrated microdevice as a multiplex immunofluorescence platform for highly sensitive and selective detection of avian influenza virus (AIV) is described. The 3D morphology and unique optical property of the ZnO nanorods boost the detection limit of the H5N2 AIV to as low as 3.6 × 10³ EID₅₀ mL^?1 (EID₅₀: 50% embryo infectious dose), which is ≈22 times more sensitive than conventional enzyme‐linked immunosorbent assay. The entire virus capture and detection process could be completed within 1.5 h with excellent selectivity. Moreover, this microfluidic biosensor is capable of detecting multiple viruses simultaneously by spatial encoding of capture antibodies. One prominent feature of the device is that the captured H5N2 AIV can be released by simply dissolving ZnO nanorods under slightly acidic environment for subsequent off‐chip analyses. As a whole, this platform provides a powerful tool for rapid detection of multiple pathogens, which may extent to the other fields for low‐cost and convenient biomarker detection.     

Measuring markers of liver function using a micropatterned paper device designed for blood from a fingerstick

This paper describes a paper-based microfluidic device that measures two enzymatic markers of liver function (alkaline phosphatase, ALP, and aspartate aminotransferase, AST) and total serum protein. A device consists of four components: (i) a top plastic sheet, (ii) a filter membrane, (iii) a patterned paper chip containing the reagents necessary for analysis, and (iv) a bottom plastic sheet. The device performs both the sample preparation (separating blood plasma from erythrocytes) and the assays; it also enables both qualitative and quantitative analysis of data. The data obtained from the paper-microfluidic devices show standard deviations in calibration runs and "spiked" standards that are acceptable for routine clinical use. This device illustrates a type of test useable for a range of assays in resource-poor settings.     

General concept of high-performance amperometric detector for microfluidic (bio)analytical chips

In this work, we established theoretically that amperometric detector arrays consisting of a series of parallel band microelectrodes placed on the wall of a microchannel may offer excellent analytical detection performances when implemented onto microfluidic (bio)analytical devices after the separative stages. In combination with the concentration imprinting strategies reported in a previous work, these exceptional performances may be extended to nonelectroactive or poorly diffusing analytes. Using an array of electrodes instead of a large single band allows the whole core of the channel to be probed though keeping an excellent time resolution. Thus, analytes with close retention times may be characterized individually with a resolution which eventually outpaces that of spectroscopic detections. Such important advantages may be obtained only through a complete understanding of the complex coupling between diffusional and convective transport of molecules in microfluidic solutions near an electrochemical detector. As a consequence, the conditions underlying the theoretical data presented in this work have been selected after optimizing procedures rooted on previous theoretical analyses. They will be fully disclosed in a series of further works that will also establish the experimental performances of such amperometric detectors and validate the present concept.     

Microfluidic electrochemical aptameric assay integrated on-chip: a potentially convenient sensing platform for the amplified and multiplex analysis of small molecules

Aptamers are artificial oligonucleotides that have been widely employed to design biosensors (i.e., aptasensors). In this work, we report a microfluidic electrochemical aptamer-based sensor (MECAS) by constructing Au-Ag dual-metal array three-electrode on-chip for multiplex detection of small molecules. In combination with the microfluidic channels covering on the glass chip, different targets are transported to the Au electrodes integrated on different positions of the chip. These electrodes are premodified by different kinds of aptamers, respectively, to fabricate different sensing interfaces which can selectively capture the corresponding target. It is an address-dependent sensing platform; thus, with the use of only one electrochemical probe, multitargets can be recognized and detected according to the readout on a corresponding aptamer-modified electrode. In the sensing strategy, the electrochemical probe, [Ru(NH(3))(6)](3+) (RuHex), which can quantitatively bind to surface-confined DNA via electrostatic interaction, was used to produce chronocoulometric signal; Au nanoparticles (AuNPs) were used to improve the sensitivity of the sensor by amplifying the detection signals. Moreover, the sensing interface fabrication, sample incubation, and electrochemical detection were all performed in microfluidic channels. By using this detection chip, we achieved the multianalysis of two model small molecules, ATP, and cocaine, in mixed samples within 40 min. The detection limit of ATP was 3 × 10(-10) M, whereas the detection limit of cocaine was 7 × 10(-8) M. This Au-Ag dual metal electrochemical chip detector integrated MECAS was simple, sensitive, and selective. Also it is similar to a dosimeter which accumulates signal upon exposure. It held promising potential for designing electrochemical devices with high throughput, high automation, and high integration in multianalysis.     

整合纸基微流控检测芯片的食药纸包装

目的探索纸质食品、药品包装与纸基微流控检测芯片的整合方法与规律。方法在传统纸质食品或药品包装的内表面,通过喷蜡打印的方法,整合具有特定生物化学检测作用的纸基微流控芯片,并探索微流体在包装内表面构成的纸基微流控芯片中的运用规律。结果通过喷蜡打印,成功地将纸基微流控芯片整合在了传统纸质食药包装的内表面,经过测试可以完成液体pH检测等基础生物化学检测应用。结论将纸基微流控芯片与食药纸质包装相结合,为食品、药品的实时和现场自我检测提供了新的思路和手段,该方法不仅成本低廉、易于操作,且检测精度高。     

Quantum dot (QD)-modified carbon tape electrodes for reproducible electrochemiluminescence (ECL) emission on a paper-based platform

Stable and sensitive electrochemiluminescence (ECL) detection relies on successful immobilization of quantum dots (QDs) on working electrodes. Herein, we report a new technique to apply double-sided carbon adhesive tape as the working electrode to improve the stability and reproducibility of QD-based ECL emission. CdS QD-modified electrodes were prepared by dropping and drying CdS QD suspension on the carbon adhesive tape supported by indium tin oxide (ITO) glass. The ECL detection was performed with the prepared electrode on a paper-based platform. We tested our system using H(2)O(2) of various concentrations and demonstrated that consistent ECL emission could be obtained. We attribute stable and reproducible ECL emission to the robust attachment of CdS QDs on the carbon adhesive tape. The proposed method could be used to quantify the concentration of dopamine from 1 μM to 10 mM based on the quenching effect of dopamine on ECL emission of CdS QD system using H(2)O(2) as the coreactant. Our approach addressed the problem in the integration of stable QD-based ECL detection with portable paper-based analytical devices. The similar design offers great potential for low-cost electrochemical and ECL analytical instruments.     

Development of biosensor based on imaging ellipsometry and biomedical applications

So far, combined with a microfluidic reactor array system, an engineering system of biosensor based on imaging ellipsometry is installed for biomedical applications, such as antibody screen, hepatitis B markers detection, cancer markers spectrum and virus recognition, etc. Furthermore, the biosensor in total internal reflection (TIR) mode has be improved by a spectroscopic light, optimization settings of polarization and low noise CCD which brings an obvious improvement of 10 time increase in the sensitivity and SNR, and 50 times lower concentration in the detection limit with a throughput of 48 independent channels and the time resolution of 0.04 S.