Encapsulation of glucose oxidase microparticles within a nanoscale layer-by-layer film: immobilization and biosensor applications (original) (raw)
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Materials Science and Engineering: C, 2006
ABSTRACT In order to determine the best kind of matrix that enables preservation of the enzymatic and electro-enzymatic activity of immobilized enzyme and provides accessibility towards substrate, various host materials (electropolymerized polypyrrole films, alginate polysaccharide, biocompatible synthetic latex and inorganic clays (laponite and layered double hydroxides LDHs), differing in permeability, ion exchange properties and hydrophobic–hydrophilic character, were compared for the fabrication of amperometric glucose biosensors. The electrochemical assays were performed by potentiostating the enzyme electrodes at 0.6 V vs. Ag/AgCl in order to oxidize the hydrogen peroxide enzymatically generated in the presence of glucose and oxygen. The highest sensitivity and maximum current density were recorded for laponite (82.3 mA M−1 cm−2 and 410 μA cm−2 respectively) and LDHs (55 mA M−1 cm−2 and 417 μA cm−2, respectively).
Improved enzyme immobilization for enhanced bioelectrocatalytic activity of glucose sensor
2009
In this work, we report the development of a highly sensitive and stable glucose sensor based on the synergetic effect of multi-walled carbon nanotubes (MWNTs) and ZnO nanoparticles. Since the isoelectric point (IEP) of glucose oxidase (GOx) is significantly different from that of ZnO nanoparticles, GOx was electrostatically bound to ZnO nanoparticles decorated onto a negatively charged MWNTs layer at the electrode surface via VLS growth. A cationic polydiallyldimethylammonium chloride (PDDA) layer was further coated onto the GOx-contained ZnO nanoparticle layer to prevent the leakage of GOx. This unique multi-layer structure (PDDA/GOx/ZnO/MWNTs) provided a favorable microenvironment to maintain the bioactivity of GOx, which led to rapid amperometric response toward glucose. By loading 0.5-U GOx at the sensor surface, we obtained a wide linear response range of 0.1-16 mM for this sensor. High sensitivity of 50.2 mA cm −2 M −1 was obtained with increased loading of GOx (2.0 U), leading to an even lower detection limit of 250 nM. This nanomaterials-based glucose sensor was highly sensitive and showed favorable stability over a relatively long-term storage (160 days). Importantly, we challenged our sensor with 100 human blood serum samples, and obtained consistent results with the classic spectrometric assay (correlation coefficient, 0.997).
The effect of the layer structure on the activity of immobilized enzymes in ultrathin films
Journal of Colloid and Interface Science, 2006
The molecular engineering capability of the layer-by-layer (LbL) method for fabricating thin films has been exploited in order to immobilize glucose oxidase (GOD) in films with alternating layers of chitosan. Chitosan was proven a good scaffolding material, as GOD molecules preserved their catalytic activity towards glucose oxidation. Using electrochemical measurements, we showed that chitosan/GOD LbL films can be used to detect glucose with a limit of detection of 0.2 mmol l −1 and an activity of 40.5 µA mmol −1 L µg −1 , which is highly sensitive when compared to other sensors in previous reports in the literature. The highest sensitivity of the LbL film was achieved when only the top layer contained GOD, thus indicating that GOD in inner layers did not contribute to glucose oxidation, probably because it hampers analyte diffusion and electron transport through the deposited layers. This may be explained by the dense packing of GOD molecules in the LbL films with chitosan, as inferred from estimates of the amount of GOD adsorbed per layer using a quartz crystal microbalance.
2013
In this thesis, the research work was focused on the immobilization of different enzymes (oxidases and dehydrogenases) into biocomposite silica matrix with the aim of amperometric biosensors construction. Then, the structured nanomaterials were introduced in the system in order to improve the characteristics of biosensors. The method of electrochemically-assisted deposition was chosen for the immobilization of enzymes on the surface of nanomaterials as it provides possibility of fine tuning of film thickness allowing covering each individual nanoobject. The feasibility of this was shown while modifying the platinum nanofibers, which demonstrate high electroactive surface and H2O2 oxidation rate, with silica-glucose oxidase biocomposite. The electrochemically-assisted deposition also allows the express modification of gold screen-printed electrodes with silica-choline oxidase biocomposite making possible the quick fabrication of cheap choline biosensors with high analytical character...
Analytical and Bioanalytical Chemistry, 2008
Protective polymer coatings have been used to enhance the retention of enzymes in sol-gel films as immobilisation phases in electrochemical biosensors. Carbon film electrodes were electrochemically modified with poly(neutral red) (PNR). These electrodes were coated with oxysilane sol-gels incorporating glucose oxidase and an outer coating of carboxylated PVC (CPVC) or polyurethane (PU), with and without Aliquat-336 or isopropyl myristate (IPM) plasticizer, was applied. The biosensors were characterised electrochemically using cyclic voltammetry and amperometry, electrochemical impedance spectroscopy and scanning electron microscopy. Impedance spectra showed that the electrode surface is most active when the sol-gel-GOx layer is not covered with a membrane. However, membranes without plasticizer extend the lifetime of the biosensor to more than 2 months when PU is used as an outer membrane. The linear range of the biosensors was found to be 0.05-0.50 mM of glucose and the biosensor with PU outer membrane exhibited higher sensitivity (ca.117 nA mM −1 ) in the region of linear response than that with CPVC. The biosensors were applied to glucose measurement in natural samples of commercial orange juice.
A novel enzyme entrapment in SU-8 microfabricated films for glucose micro-biosensors
Biosensors and Bioelectronics, 2010
The present work investigates the utilisation of the widely used SU-8 photoresist as an immobilisation matrix for glucose oxidase (GOx) for the development of glucose microbiosensors. The strong advantage of the proposed approach is the simultaneous enzyme entrapment during the microfabrication process within a single step, which is of high importance for the simplification of the BioMEMS procedures. Successful encapsulation of the enzyme GOx in "customised" SU-8 microfabricated structures was achieved through optimisation of the one-step microfabrication process. Although the process involved contact with organic solvents, UV-light exposure, heating for pre-and post-bake and enzyme entrapment in a hard and rigid epoxy resin matrix, the enzyme retained its activity after encapsulation in SU-8. Measurements of the immobilised enzyme's activity inside the SU-8 matrix, were carried out using amperometric detection of hydrogen peroxide in a 3-electrode setup. Films without enzyme showed negligible variation in current upon the addition of glucose, as opposed to films with encapsulated enzyme which showed a very clear increase in current. Experiments using films of increased thickness or enzyme concentration, showed a higher response, thus proving that the enzyme remained active not only on the film's surface, but inside the matrix as well. The proposed enzyme immobilisation in SU-8 films opens up new possibilities for combining BioMEMS with biosensors and organic electronics.
Miniaturized thin-film biosensors using covalently immobilized glucose oxidase
Biosensors and Bioelectronics, 1991
Ah&rack The production of a miniaturized glucose sensor by means of thinfilm technology is reported. Two main problems related to miniatu~~tion and device integration were solved: (1) the microminiaturization of a suitable electrochemical cell; (2) localized enzyme immobilization with a technology well suited for device integration. The well-known glucose oxidase/H* OZ system was used to determine the glucose concentration. A miniaturized four-electrode arrangement was introduced to measure HZ& produced by the enzyme. A double working electrode array for reproducibility tests or differential measurements to suppress interferences is easily produced and can be placed on glass or flexible polymer substrates by means of thin-film technology. The enzyme was covalently coupled to a derivatized platinum thin-film working electrode by means of 12-arenequinones, which yield highly reproducible, fast and stable sensors. Measurement of a drop (5 ~1) of physiologi~l glucose solution is easily performed, giving a stable response after 40 s.
Bioelectrochemistry, 2014
Multi-wall carbon nanotubes (MWNTs) functionalized with amino groups were prepared via silane treatment using 3-aminopropyltrimethoxysilane (APS) as a silane-coupling agent. The resultant amino terminated MWNTs (AMWNTs) were applied to construct glucose biosensors with IO 4 −-oxidized glucose oxidase (IO 4 −-oxidized GOx) through the layer-by-layer (LBL) covalent self-assembly method without any cross-linker. Scanning electron microscopy (SEM) indicated that the assembled AMWNTs were almost in a form of small bundles or single nanotubes, and the surface density increased uniformly with the number of GOx/AMWNTs bilayers. From the analysis of voltammetric signals, a linear increment of the coverage of GOx per bilayer was estimated. The resulting biosensor showed excellent catalytic activity towards the electroreduction of dissolved oxygen at low overvoltage, based on which glucose concentration was monitored conveniently. The enzyme electrode exhibited good electrocatalytic response towards the glucose and that response increased with the number of GOx/AMWNTs bilayers, suggesting that the analytical performance such as sensitivity and detection limit of the glucose biosensors could be tuned to the desired level by adjusting the number of deposited GOx/AMWNTs bilayers. The biosensor constructed with four bilayers of GOx/AMWNTs showed high sensitivity of 7.46 A mM −1 cm −2 and the detection limit of 8.0 M, with a fast response less than 10 s. Because of relative low applied potential, the interference from other electro-oxidizable compounds was minimized, which improved the selectivity of the biosensors. Furthermore, the obtained enzyme electrodes also showed remarkable stability due to the covalent interaction between the GOx and AMWNTs.