John Njagi

Amperometric detection of dopamine in vivo with an enzyme based carbon fiber microbiosensor.

We developed a novel implantable enzyme-based carbon fiber biosensor for in vivo monitoring of dopamine. The biosensor is fabricated using tyrosinase immobilized in a biocompatible matrix consisting of a biopolymer, chitosan and ceria-based metal oxides, deposited onto the surface of a carbon fiber microelectrode with a diameter of approximately 100 microm. Tyrosinase catalyzes the conversion of dopamine to o-dopaquinone, and the reduction of o-dopaquinone, which requires a low potential difference, was detected electrochemically. The role of each component in the sensing layer was systematically investigated in relation to the analytical performance of the biosensor. In its optimal configuration, the biosensor demonstrated a detection limit of 1 nM dopamine, a linear range of 5 orders of magnitude between 10 nM and 220 microM, a sensitivity of 14.2 nA x microM(-1), and good selectivity against ascorbic acid, uric acid, serotonin, norepinephrine, epinephrine, and 3,4-dihydroxy-l-phenylalanine (L-DOPA). The system provided continuous, real time monitoring of electrically stimulated dopamine release in the brain of an anesthetized rat. Levels of dopamine up to 1.69 microM were measured. This new implantable dopamine biosensor provides an alternative to fast scan cyclic voltammetry for in vivo monitoring of dopamine.

Mixed Ceria-Based Metal Oxides Biosensor for Operation in Oxygen Restrictive Environments

The unique catalytic, electrochemical, and oxygen storage properties of ceria and mixed ceria/titania hybrid composites were used to fabricate a new type of electrochemical enzyme biosensor. These materials provided increased analytical performance and possibilities for operation in oxygen-free conditions of an oxidase enzyme biosensor using tyrosinase as a model example. The investigation of the enzymatic reaction in the presence and absence of oxygen was first carried out using cyclic voltammetry. The results were used to identify the role of each metal oxide in the immobilization matrix and fabricate a simple amperometric tyrosinase biosensor for the detection of phenol and dopamine. The biosensor was optimized and characterized with respect to response time, detection limit, linear concentration range, sensitivity, and kinetic parameters. The detection limit for phenol was in the nanomolar range, with a detection limit of 9.0 x 10(-9) M and a sensitivity of 86 mA M(-1) in the presence of oxygen and of 5.6 x 10(-9) M and a sensitivity of 65 mA M(-1) in the absence of oxygen. The optimized biosensor also showed selective determination of the neurotransmitter dopamine with a detection limit of 3.4 x 10(-8) M and a sensitivity of 14.9 mA M(-1) in the presence of oxygen and of 4.2 x 10(-8) M and 14.8 mA M(-1) in the absence of oxygen. This strategy shows promise for increasing the sensitivity of oxidase enzyme sensors and provides opportunities for operation in oxygen limited conditions. It can also be extended for the development of other enzyme biosensors.

Electrochemical Studies of Ceria as Electrode Material for Sensing and Biosensing Applications

This work presents electrochemical studies of ceria as an electrode material and describes new possibilities of exploring the unique catalytic and redox properties of this material for the development of a simple enzymeless sensor for the determination of H 2 O 2 and for biosensor manufacturing. The investigation of the electrocatalytic activity of ceria toward the oxidation and reduction of hydrogen peroxide was first carried out using cyclic voltammetry in alkaline and neutral conditions. A possible reaction mechanism was suggested involving different cerium oxidation states and reactive species. The ceria-based sensor resulted in rapid, sensitive, and accurate measurement of H 2 O 2 , with a response time of less than 10 s, a detection limit of 98 nM, and a sensitivity of 0.24 μA μM -1 . This material provided a biocompatible matrix for the immobilization of glucose oxidase and facilitated direct electron transfer between its active center and the electrode surface, with an electron-transfer rate constant of 18.3 s -1 and a surface coverage of 1.4 × 10 -11 mol/cm 2 . This study introduces ceria as electrode material for sensing and biosensing applications.

Electrochemical Quantification of Serotonin in the Live Embryonic Zebrafish Intestine

We monitored real-time in vivo levels of serotonin release in the digestive system of intact zebrafish embryos during early development (5 days postfertilization, dpf) using differential pulse voltammetry with implanted carbon fiber microelectrodes modified with carbon nanotubes dispersed in nafion. A detection limit of 1 nM, a linear range between 5 and 200 nM, and a sensitivity of 83.65 nA x microM(-1) were recorded. The microelectrodes were implanted at various locations in the intestine of zebrafish embryos. Serotonin levels of up to 29.9 (+/-1.13) nM were measured in vivo in normal physiological conditions. Measurements were performed in intact live embryos without additional perturbation beyond electrode insertion. The sensor was able to quantify pharmacological alterations in serotonin release and provide the longitudinal distribution of this neurotransmitter along the intestine with high spatial resolution. In the presence of fluvoxamine, a selective serotonin reuptake inhibitor (SSRI), concentrations of 54.1 (+/-1.05) nM were recorded while in the presence of p-chloro-phenylalanine (PCPA), a tryptophan hydroxylase inhibitor, the serotonin levels decreased to 7.2 (+/-0.45) nM. The variation of serotonin levels was correlated with immunohistochemical analysis. We have demonstrated the first use of electrochemical microsensors for in vivo monitoring of intestinal serotonin levels in intact zebrafish embryos.

Deposition of continuous platinum shells on gold nanoparticles by chemical precipitation

Continuous platinum shells consisting of ~5 atomic layers were deposited onto preformed gold seeds in aqueous medium by reducing hexachloroplatinic acid with ascorbic acid. By controlling the reduction kinetics of Pt(IV) species and the properties of the substrate, it was possible to ensure a slow and controlled deposition of platinum atoms onto the gold cores. Electrochemical evaluations revealed the presence of a compact platinum shell. The mass specific oxygen reduction activity of platinum in the AuPt core-shell nanoparticles was found to be four times higher than that of platinum black and comparable to that of polycrystalline bulk metal.

Nanostructured materials for enzyme immobilization and biosensors

Publisher Summary The unique electrical, optical, catalytic, and magnetic properties of materials in which the structural elements are in the nanometer size (broadly defined as nanostructured materials) have attracted considerable interest for designing powerful enzyme-based biocatalytic systems. This chapter discusses recent advances in the study and use of various classes of nanostructured materials (carbon nanotubes, nanofibers, and nanowires, nanoparticles and nanocrystals, mesoporos silica and, composite materials) for enzyme immobilization and describes selected examples of their application in the development of biologically active systems and biosensors. Different strategies used for the functionalization of nanostructures with enzymes, development and characterization of the nanostructure assembly, evaluation of biological activity and generation of response and sensing function are also discussed.

Nitrilotriacetic acid: A novel reducing agent for synthesizing colloidal gold

We report for the first time that nitrilotriacetic acid (NTA) is an effective reductant for the preparation of stable dispersions of uniform gold nanoparticles. The method described is capable of generating stable sols with a metal concentration as high as 1.5×10(-3)moldm(-3). The size of gold nanoparticles can be tuned from 10 to 160 nm by adjusting the stoichiometric excess of NTA. For a constant [Au]/[NTA] ratio the temperature affects the reduction kinetics but has little impact on the size of gold nanoparticles. The mechanisms of the reduction of Au(III) species and the formation and stabilization of gold nanoparticles are discussed.

Deposition of continuous nickel shells on polymer microspheres.

Anisotropic conductive adhesives (ACAs) are widely used as interconnect materials in the manufacturing of LCD screens. To be integrated in a broader range of applications, several technical and economical issues still need to be addressed. Encapsulating the polymer particles within continuous, compact, and adhering metallic conductive shells is one of these challenges. This work describes a method for depositing nickel layers with different thickness (30-120nm) onto monosized polymer particles. The novelty of the approach consists in modifying the surface of polymer particles with linear polymeric amines. We show that by increasing amine chain length the structure and adhesion of deposited nickel shell are significantly improved. The effect of key parameters of the electroless Ni plating are discussed and illustrated.

Electrical four-point probing of spherical metallic thin films coated onto micron sized polymer particles

Micron-sized metal-coated polymer spheres are frequently used as filler particles in conductive composites for electronic interconnects. However, the intrinsic electrical resistivity of the spherical thin films has not been attainable due to deficiency in methods that eliminate the effect of contact resistance. In this work, a four-point probing method using vacuum compatible piezo-actuated micro robots was developed to directly investigate the electric properties of individual silver-coated spheres under real-time observation in a scanning electron microscope. Poly(methyl methacrylate) spheres with a diameter of 30 μm and four different film thicknesses (270 nm, 150 nm, 100 nm, and 60 nm) were investigated. By multiplying the experimental results with geometrical correction factors obtained using finite element models, the resistivities of the thin films were estimated for the four thicknesses. These were higher than the resistivity of bulk silver.

Detection and prediction of concentrations of neurotransmitters using voltammetry and pattern recognition

Neurotransmitters (NTs) are substances in the brain which are responsible for the transmission of neurological impulses. Changes in their concentrations are associated with numerous behavioral and physiological processes and neurological disorders. As opposed to the traditional chromatographic and capillary electrophoresis, using electrochemical sensors is a fast and inexpensive way to determine concentrations of NTs. In this study we measure the combination of dopamine (DA) and serotonin (SE) with glassy carbon electrodes and differential pulse voltammetry. The major challenge using this method is to differentiate between different NTs, since the signal obtained from the electrode represents the interactive effect of both NTs present. We address this problem through methods of pattern recognition which relate the voltammetric measurements provided by the sensor to the concentration of individual NTs. Two methods of pattern recognition were applied (PCR and PLS-regression). The best rates of correct classification for the validation sets ranged at 42–62% (DA) and 33–50% (SE). When the ranges for correct prediction were extended to include one level above and below the true concentration level, the rates values ranged at 81–91% (DA) and 91–100%(SE). These findings suggest that pattern recognition can be used to model the interaction between different neurotransmitters to predict actual concentrations of neurotransmitters using voltammetry.