Róbert E. Gyurcsányi

Microfabricated ISEs: critical comparison of inherently conducting polymer and hydrogel based inner contacts

A rigorous side by side comparison of miniature planar potassium-selective electrodes with hydrogel and potassium hexacyanoferrate(II)/(III) doped polypyrrole (PPy/FeCN) based inner contacts is presented. The planar electrodes were manufactured by screen printing as four- and five-site arrays on ceramic substrates. These electrode arrays were incorporated into a flow-through cell, which could accommodate nine electrode sites. Two identical flow cells were connected in series and the effect of the inner contacts on the analytical performance of the respective electrodes has been critically evaluated. The time necessary to reach steady state conditions has been determined and the effect of experimental parameters (temperature, ambient light intensity, CO(2), and O(2) concentration of the sample) on the potential stability of the electrodes was analyzed. At controlled temperature, the drift of the planar potassium electrodes with hydrogel and PPy/FeCN solid contact were 0.11+/-0.02mVh(-1) and 0.03+/-0.007mVh(-1), respectively. The experimental data proved that there is no aqueous film formation between the PPy/FeCN film and the potassium-selective solvent polymeric membrane.

Novel polypyrrole based all-solid-state potassium-selective microelectrodes

Potassium-selective potentiometric microelectrodes with a polypyrrole solid internal contact were fabricated by the application of a potassium-selective bis-crown ether ligand based, plasticized poly(vinyl chloride) (PVC) membrane to the surface of conducting polymer modified Pt, Au or C micro disk electrodes. The selectivity and sensitivity of the new type of potentiometric potassium microelectrode were found to be comparable with those of the conventional macro ion-selective electrodes and of the micropipet type microelectrodes based on the same ionophore. The ease of preparation and robustness are the main advantages of this new electrode design, which can replace the classical micropipet type microelectrodes in many applications. The microelectrodes showed good dynamic characteristics and were used successfully in a wall-jet cell incorporated flow injection analysis system and as a measuring tip in scanning electrochemical microscopy. The polypyrrole modified substrate electrodes can be made selective for different ions just by changing the ionophore in the plasticized PVC membrane, which can extend their use to a wide range of applications.

Amperometric microcells for alkaline phosphatase assay

To develop simple electrochemical immunoassays, a screen printed amperometric microcell with graphite working and Ag/AgCl reference electrodes was tested for the determination of alkaline phosphatase enzyme (ALP) and anti-humanIgG conjugated ALP (alpha-hIgG-ALP) activity in 5-10 microl samples. To ensure reproducible, steady state conditions, the working electrode surface was coated with mass-transport controlling hydrogel layer. The kinetic response curves of the hydrogel coated electrodes were linear. In addition, the hydrogel layer reduced the nonspecific adsorption of the alpha-hIgG-ALP conjugate on the working electrode surface. The measurements were made in the range of 2 divided by 4000 mU ml(-1) enzyme activities using ascorbic acid 2-phosphate (AAP) as the enzyme substrate. AAP is commercially available, non-toxic and has excellent stability. The sensitivity of the determinations was about 71% of the sensitivity which could be achieved using p-aminophenylphosphate (PAPP), a not easily accessible and unstable enzyme substrate. The experimentally determined kinetic parameters of the ALP enzyme catalyzed reactions were the same with the bare and hydrogel layer coated electrodes.

Aptamer-based biochips for label-free detection of plant virus coat proteins by SPR imaging.

Specific detection of virus strains by affinity-based bioassays is often limited by the availability of ligands able to differentiate among close homologues of virus coat proteins. As viruses are prone to mutation, the ligand generation should, in addition, be fast enough to allow rapid identification of new varieties. These two criteria are difficult to be fulfilled by antibodies; however, they open up opportunities for aptamer-based detection. Here we report on the feasibility of selectively detecting the apple stem pitting virus (ASPV) coat proteins (PSA-H, MT32) using original DNA aptamers. Surface plasmon resonance (SPR) imaging was used together with aptamer-modified sensor chips to optimize the aptamer immobilization for highest sensitivity and to characterize the aptamer-virus coat protein binding. Different parameters affecting this binding, such as the aptamer flanking, surface coverage, and type of spacer molecules, were identified and their influence was determined. A direct label-free method is proposed for assessing the ASPV based on the detection of the respective virus coat proteins in plant extracts.

Polyaniline nanoparticle-based solid-contact silicone rubber ion-selective electrodes for ultratrace measurements.

Silicone rubber (SR)-based solid-contact ion-selective electrodes (ISEs) have been prepared for the first time with an electrically conducting polymer as the solid-contact (SC) layer. The Ca(2+)- and Ag(+)-selective electrodes were based on the ionophores ETH 1001 and o-xylylenebis(N,N-diisobutyl dithiocarbamate), respectively, integrated in room temperature vulcanizing silicone rubber (RTV 3140). The SC consisted of a polyaniline nanoparticle dispersion, which was found to considerably lower the impedance of the SCISEs in comparison to the SR-based coated wire electrodes (CWE). For the CaSCISEs, the bulk membrane resistance decreased from 700 MΩ (CaCWE) to 35 MΩ. Both the Ca(2+)- and Ag(+)-selective SCISEs exhibited nanomolar detection limits with fast Nernstian responses down to 10(-8) M. The potential response of the SCISEs was not influenced by light. The selectivities of the CaSCISEs were similar and for the AgSCISE better than their plasticized PVC-based analogues. Thus, SR seems to be a viable alternative to PVC membranes in ISE applications that require low water uptake, good adhesion, and robust and fast potential responses at submicromolar sample concentrations.

Hyphenated FT-IR-attenuated total reflection and electrochemical impedance spectroscopy technique to study the water uptake and potential stability of polymeric solid-contact ion-selective electrodes.

A new hyphenated method utilizing FT-IR-attenuated total reflection (ATR) and electrochemical impedance spectroscopy (EIS) is presented to correlate the water uptake with concomitant potential and impedance changes of polymeric coated-wire electrodes (CWEs) and solid-contact ion-selective electrodes (SCISEs). The Ca(2+)-selective silicone rubber (RTV 3140) based SCISEs with poly(3-octylthiophene) (POT) as the solid-contact (SC) showed good correlation between a very low water content at the Pt-coated ZnSe substrate/SC interface and a superior potential stability. This is due to the hydrophobicity of both RTV 3140 and POT and the approximately 2 orders of magnitude lower water diffusion coefficients in POT compared to RTV 3140. Practically no potential drift could be observed during 24 h when unconditioned CaSCISEs were contacted with 10(-3) M CaCl(2), in contrast to the Ca(2+)-selective CWEs with considerably higher water uptake and potential drift. The CaSCISEs had a fast Nernstian response with a detection limit of 8 × 10(-9) M Ca(2+) and a good reproducibility and stability of the standard potential, which indicates that the CaSCISEs does not require any conditioning prior to use.

Biorecognition-modulated ion fluxes through functionalized gold nanotubules as a novel label-free biosensing approach

A novel biosensing principle is presented, based on the potentiometric monitoring of an indicator ion such as Ca2+, whose zero-current flux through chemically modified nanochannels is altered by biorecognition events.

Electrochemical methods for the determination of the diffusion coefficient of ionophores and ionophore–ion complexes in plasticized PVC membranes

The diffusion coefficients of active components in ion-selective membranes have a decisive influence on the life-time and detection limit of the respective ion-selective electrodes, as well as influencing the rate of polarization and relaxation processes of electrically perturbed ion sensors. Therefore, the rational design of mass transport controlled ion-selective electrodes with sub-nanomolar detection limits requires reliable data on the diffusion coefficients. We have implemented electrochemical methods for the quantitative assessment of both the diffusion coefficients of free ionophores and ion-ionophore complexes. The diffusion coefficients of the pH-sensitive chromoionophore ETH 5294 and the calcium-selective ionophore ETH 5234 were determined in plasticized PVC membranes with different PVC to plasticizer ratios. The diffusion coefficient of the free chromoionophore determined by a chronoamperometric method was validated with optical methods for a variety of membrane compositions. The calcium-selective ionophore ETH 5234 was used as a model compound to assess the diffusion coefficient of the ion-ionophore complex calculated from the time required for the complexes to cross a freshly prepared membrane during potentiometric ion-breakthrough experiments. The difference between the diffusion coefficients of the free ionophore ETH 5234 and the ion-ionophore complex was found to be significant and correlated well with the geometry of the respective species.

FTIR-ATR Study of Water Uptake and Diffusion through Ion-Selective Membranes Based on Poly(acrylates) and Silicone Rubber

For the first time, FTIR-ATR spectroscopy was used to study the water uptake and its diffusion in ion-selective membranes (ISMs) based on poly(acrylates) (PAs) and silicone rubber (SR), which are emerging materials for the fabrication of ISMs for ultratrace analysis. Three different types of PA membranes were studied, consisting of copolymers of methyl methacrylate with n-butyl acrylate, decyl methacrylate, or isodecyl acrylate. Numerical simulations with the finite difference method showed that in most cases the water uptake of the PA and SR membranes could be described with a model consisting of two diffusion coefficients. The diffusion coefficients of the PA membranes were approximately 1 order of magnitude lower than those of plasticized poly(vinyl chloride) (PVC)-based ISMs and only slightly influenced by the membrane matrix composition. However, the simulations indicated that during longer contact times, the water uptake of PA membranes was considerably higher than that for plasticized PVC membranes. Although the diffusion coefficients of the SR and plasticized PVC membranes were similar, the SR membranes had the lowest water uptake of all membranes. This can be beneficial in preventing the formation of detrimental water layers in all-solid-state ion-selective electrodes. With FTIR-ATR, one can monitor the accumulation of different forms of water, i.e., monomeric, dimeric, clustered, and bulk water.

Is less more? Lessons from aptamer selection strategies

Aptamers have many inherent advantages originating from their in vitro selection and tailored chemical synthesis that makes them appealing alternatives of antibodies in bioaffinity assays. However, what ultimately matters, and that is the prerequisite to give way to all these advantages, is how well, and how selectively the aptamers bind to their targets. With the aptamer selection largely in the hand of life scientists, analytical chemists focused mostly on methodological development of aptamer-based assays using a fairly restricted number of aptamers to prove their concepts. However, ideally the development of an aptamer-based assay should start from the selection of aptamers to ensure their proper functionality in real samples. For instance information on the sample matrix can be implemented within counter-selection steps to discard aptamer candidates that show cross-reactivity to matrix components or critical interferents. In general, a larger consideration of the analytical use during selection and characterization of aptamers have been shown to increase the applicability of aptamers. Therefore, this review is a short, subjective view on trends in aptamer development highlighting factors to consider during their selection for a successful analytical application.