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Amperometry

Amperometry is an electrochemical technique that measures the current produced by the oxidation or reduction of an electroactive species at an electrode under a constant applied potential, yielding a signal directly proportional to the analyte's concentration in a diffusion-controlled process.^[1] In amperometric measurements, a three-electrode system is typically employed, consisting of a working electrode (often platinum or gold), a reference electrode (such as Ag/AgCl), and a counter electrode to complete the circuit and maintain the desired potential at the working electrode.^[2] The applied potential drives the redox reaction of the target species, generating a faradaic current that reflects the rate of electron transfer, governed by principles like the Cottrell equation for chronoamperometry, where current decreases with the square root of time due to diffusion limitations: I = nF A C \sqrt{\frac{D}{\pi t}}, with n as the number of electrons, F as Faraday's constant, A as electrode area, C as bulk concentration, D as diffusion coefficient, and t as time.^[3] This method distinguishes itself from voltammetry by holding the potential constant rather than sweeping it, enabling steady-state or transient current monitoring for quantitative analysis.^[4] The foundations of amperometry trace back to early 20th-century electrochemistry, but its analytical prominence emerged in the mid-20th century with advancements in electrode design and instrumentation, including the development of the dropping mercury electrode by Heyrovský in the 1920s, which influenced polarographic techniques closely related to amperometry.^[5] A pivotal milestone occurred in the 1950s–1960s through Leland C. Clark's invention of the oxygen electrode in 1956 and the first enzyme-based amperometric biosensor for glucose in 1962, which revolutionized biosensing by coupling enzymatic reactions with current detection.^[6] Amperometry finds extensive applications in analytical chemistry, particularly in clinical diagnostics for monitoring biomolecules like glucose, lactate, and uric acid via portable biosensors; environmental analysis for detecting pollutants such as heavy metals and phenols; and industrial processes including gas sensing for oxygen or toxic gases in electrolytic cells.^[2] Its advantages—high sensitivity (down to nanomolar levels), selectivity through potential control, rapid response times, and compatibility with miniaturization—make it ideal for point-of-care devices, flow-injection analysis, and chromatography detectors, though challenges like electrode fouling require strategies such as pulsed potentials or mediator use.^[1]

Fundamentals

Definition and Scope

Amperometry is an electrochemical technique that involves the measurement of electric current resulting from the electrochemical oxidation or reduction of an analyte species at an electrode surface, where the electrode potential is maintained at a constant value.^[7] This current is directly proportional to the concentration of the electroactive species in the solution, enabling quantitative analysis. The method typically employs a three-electrode system, consisting of a working electrode (where the reaction occurs), a reference electrode (to control potential), and a counter electrode (to complete the circuit).^[7] The scope of amperometry encompasses its primary application in the quantitative determination of analytes in solution, particularly for electroactive compounds such as neurotransmitters, oxygen, and carbohydrates.^[8] It forms a subset of voltammetric techniques but is distinguished by its emphasis on monitoring time-dependent current responses at a fixed potential, rather than sweeping the potential to generate voltammograms.^[7] This fixed-potential approach allows for both steady-state measurements, where current stabilizes after diffusion-limited conditions are reached, and transient measurements capturing dynamic processes like reaction kinetics. In comparison to related methods, amperometry differs from polarography, which utilizes a dropping mercury electrode to minimize surface effects and provide renewable electrode surfaces, whereas amperometry commonly employs solid electrodes like platinum or carbon for more stable and versatile operation.^[9] Similarly, it contrasts with general voltammetry, where the applied potential is varied linearly or in steps to probe electrochemical behavior across a range, as opposed to amperometry's constant potential for selective, concentration-focused detection.^[7] The technique traces its origins to foundational work in electrochemistry from the early 20th century, laying the groundwork for modern analytical applications.^[10]

Electrochemical Basis

Amperometry relies on the fundamental electrochemical process of redox reactions, where an analyte species undergoes oxidation or reduction at the surface of the working electrode upon the application of a suitable potential. In this process, electrons are transferred between the electrode and the analyte, generating a measurable current that is proportional to the analyte concentration. For instance, the oxidation of a species like ferrocyanide (Fe(CN)₆⁴⁻) to ferricyanide (Fe(CN)₆³⁻) occurs at an anodic potential, while reduction reactions, such as the conversion of oxygen to water, take place at cathodic potentials.^[11]^[12] The electrochemical setup typically employs a three-electrode system to precisely control the potential and measure the current. The working electrode serves as the site where the redox reaction occurs, often constructed from materials like platinum or carbon to provide a stable surface for electron transfer. The reference electrode, such as the saturated calomel electrode (SCE) with a fixed potential of +0.244 V versus the standard hydrogen electrode, maintains a constant reference point to accurately set the working electrode potential without interference from solution resistance. The counter (or auxiliary) electrode, usually a platinum wire, completes the electrical circuit by facilitating current flow and balancing the charge, preventing potential shifts at the working electrode due to ohmic losses.^[11]^[12] At the electrode-solution interface, several phenomena govern the current response. The electrical double layer forms due to charge separation, with ions accumulating to counter the electrode's charge, creating a capacitor-like structure that influences ion distribution near the surface. Charge transfer kinetics determine the rate of electron exchange between the electrode and analyte; reversible systems exhibit fast kinetics, allowing equilibrium to be maintained, while irreversible ones show slower rates that can limit the observed current. Upon potential application, an initial transient current spike arises from the charging of this double layer, which rapidly decays as the faradaic (redox-related) current dominates.^[11]^[12] Analyte delivery to the electrode surface is controlled by mass transport mechanisms, including diffusion, migration, and convection, which collectively determine the steady-state current. Diffusion, driven by concentration gradients, is the primary mode in quiescent solutions, leading to a diffusion-limited steady-state current where the rate of analyte arrival matches its consumption at the electrode. Migration, the movement of charged species under the electric field, can enhance or hinder transport depending on the analyte's charge relative to the applied potential, though it is often minimized using supporting electrolytes. Convection, induced by stirring or natural flow, thickens or thins the diffusion layer (typically 1–10 μm), accelerating mass transport and increasing the limiting current in practical setups.^[11]^[12]

Historical Development

Origins in Electrochemistry

The foundations of amperometry lie in the early principles of electrochemistry established during the 19th century, particularly Michael Faraday's laws of electrolysis published in 1833 and 1834, which quantitatively related the amount of substance altered at an electrode to the quantity of electricity passed through the electrolyte. These laws provided the groundwork for understanding current flow in electrochemical systems, emphasizing that the mass deposited or liberated is proportional to the charge and inversely proportional to the ion's equivalent weight. Building on this, Walther Nernst's diffusion layer theory in 1904 introduced the concept of a stagnant boundary layer adjacent to the electrode surface, where mass transport is governed by diffusion, laying the theoretical basis for current measurements limited by analyte diffusion to the electrode. In the early 20th century, amperometry emerged as a distinct technique from the broader field of voltammetry, with Jaroslav Heyrovský's invention of polarography in 1922 serving as a key precursor; this method used a dropping mercury electrode to record current-potential curves, enabling quantitative analysis based on diffusion-controlled currents.^[13] Heyrovský's work highlighted the analytical potential of measuring faradaic currents at controlled potentials, inspiring adaptations for constant-potential measurements. Izaak Maurits Kolthoff advanced these ideas in the 1930s through his development of amperometric titrations, where current is monitored at a fixed potential during titration to detect endpoints via changes in diffusion-limited currents, often using solid electrodes like platinum to avoid mercury's limitations.^[14] Initial applications of amperometry focused on oxygen sensing in the 1930s and 1940s, leveraging polarographic principles to measure dissolved oxygen via its reduction current at noble metal electrodes, which proved valuable for environmental and biological analyses.^[15] A pivotal device was Leland Clark's membrane-covered oxygen electrode, conceived and constructed in 1954, which enclosed a platinum cathode and silver anode behind a polyethylene membrane to selectively measure oxygen tension through amperometric reduction, marking an early practical amperometric sensor for clinical use.^[16] Instrumentation for amperometry evolved from manual potential control using galvanometers and simple cells in the early decades to more precise electronic systems in the 1940s, exemplified by Archie Hickling's invention of the three-electrode potentiostat in 1942, which employed negative feedback to maintain constant electrode potential independent of solution resistance.^[17] This transition enabled reliable constant-potential amperometry, facilitating reproducible current measurements essential for quantitative analysis up to the mid-20th century.

Key Milestones and Advances

In the 1960s, a pivotal advancement in amperometry occurred with the development of the first enzyme-based glucose biosensor by Leland C. Clark Jr., who combined glucose oxidase with an amperometric oxygen electrode to enable selective detection of glucose through enzymatic oxidation and subsequent measurement of the resulting current.^[18] This innovation laid the groundwork for modern biosensors by demonstrating how amperometry could be integrated with biological recognition elements for specific analyte detection.^[19] Commercialization followed in the 1970s, with the first glucose analyzer based on Clark's design introduced in 1975 by Yellow Springs Instrument Company, marking the transition of amperometric biosensors from laboratory prototypes to practical analytical tools.^[20] The 1980s brought significant innovations in pulsed amperometric detection (PAD), first reported by Dennis C. Johnson and colleagues in 1981, who applied potential waveforms to noble metal electrodes to detect carbohydrates by mitigating surface fouling through periodic cleaning pulses.^[21] This technique enhanced sensitivity and reproducibility for oxidizable analytes like sugars, enabling direct detection without derivatization.^[22] Concurrently, Dionex Corporation commercialized high-performance anion-exchange chromatography coupled with PAD (HPAEC-PAD) in the mid-1980s, revolutionizing anion and carbohydrate analysis by providing robust, automated detection for complex samples.^[23] Advances in microelectrodes during this decade, including carbon fiber designs introduced for in vivo amperometry, improved spatial resolution and reduced interference, facilitating applications in neuroscience and environmental monitoring.^[10] Flow-injection analysis (FIA) with amperometric detection also gained traction in the 1980s, building on FIA's invention in 1975 to enable rapid, automated sample processing with electrochemical readout for pharmaceuticals and ions. In the 1990s, the International Union of Pure and Applied Chemistry (IUPAC) played a key role in standardizing amperometric protocols and nomenclature through recommendations on electroanalytical terms, including definitions for amperometry, which promoted consistent methodology across research and industry.^[24] From the 2000s onward, amperometry integrated with microfluidics and nanomaterials expanded its scope, with carbon nanotubes and nanoparticles enhancing electron transfer and sensitivity in miniaturized devices for point-of-care diagnostics.^[25] Microfluidic systems incorporating amperometric detection emerged around 2005, enabling low-volume, high-throughput analysis in lab-on-a-chip formats for biomarkers and pollutants.^[26] Recent enhancements up to 2025 have focused on screen-printed electrodes (SPEs) for portable amperometric sensors, with modifications like nanoparticle doping improving stability and enabling smartphone-integrated devices for on-site glucose and drug monitoring.^[27] These SPE-based advances, highlighted in reviews from 2020–2024, have democratized amperometry by reducing costs and enhancing portability for field applications.^[28]

Theoretical Principles

Mechanism of Current Measurement

In amperometry, a fixed potential is applied to the working electrode, selected based on the redox potential of the target analyte to drive its oxidation or reduction, thereby generating a Faradaic current proportional to the rate of electron transfer at the electrode surface.^[29]^[30] This Faradaic current arises from the electrochemical reaction where the analyte species undergo electron exchange, distinguishing it from non-Faradaic (capacitive) contributions.^[31] The current-time profile in amperometric measurements typically begins with a rapid decay of the initial capacitive current, which charges the electrical double layer at the electrode-solution interface, followed by a diffusion-controlled Faradaic current that either reaches a steady state under convective conditions or exhibits a Cottrell-like decay in transient setups without stirring.^[32]^[33] In steady-state amperometry, such as with solution stirring or membrane diffusion, the current stabilizes as the analyte supply balances consumption at the electrode, providing a reliable signal for quantification.^[29] Detection limits in amperometry are primarily governed by the signal-to-noise ratio of the Faradaic current, where noise from background processes like residual capacitive currents or interferences can mask low analyte concentrations.^[30] Electrode materials, such as platinum or carbon, influence this ratio by affecting surface reactivity, catalytic efficiency, and fouling resistance; for instance, platinum electrodes often yield lower detection limits due to their high electrocatalytic activity for many redox species.^[30] Solution stirring enhances mass transport, reducing diffusion layer thickness and improving sensitivity, though excessive convection may introduce noise.^[29] Instrumentation for amperometric current measurement relies on a potentiostat, which applies the constant potential between the working and reference electrodes while monitoring the resulting current flow to the auxiliary electrode.^[33] The potentiostat uses operational amplifiers to maintain precise voltage control and transduce the current signal, often amplifying nanoampere-level responses for data acquisition systems that record time-dependent profiles in chronoamperometry.^[33] This setup ensures stable operation, with the acquired current data processed to extract analyte information while minimizing artifacts from uncompensated resistance or capacitance.^[29]

Mathematical Models

The mathematical models underlying amperometry provide a quantitative description of the current response as a function of time, potential, and mass transport processes. These models are derived primarily from the principles of diffusion, convection, and electrochemical thermodynamics, enabling the prediction of signal behavior under controlled conditions. Central to amperometry is the relationship between the measured faradaic current and the concentration of the electroactive species, governed by Fick's laws of diffusion and the Nernst equation for interfacial equilibria.^[34] In transient amperometry, where a constant potential is applied and the current decays over time due to diffusion limitation, the Cottrell equation describes the diffusion-controlled current for a planar electrode under semi-infinite linear diffusion conditions. The equation is given by

i(t) = \frac{n F A D^{1/2} C}{\pi^{1/2} t^{1/2}},

where i(t) is the current at time t, n is the number of electrons transferred, F is the Faraday constant, A is the electrode area, D is the diffusion coefficient, and C is the bulk concentration of the electroactive species.^[34] This form arises from solving Fick's second law of diffusion, \frac{\partial C}{\partial t} = D \frac{\partial^2 C}{\partial x^2}, with the initial condition C(x,0) = C for x > 0 and boundary conditions C(0,t) = 0 (due to rapid electroreduction at the applied potential) and C(\infty,t) = C. The solution for the concentration profile is C(x,t) = C \left[1 - \erf\left(\frac{x}{2\sqrt{Dt}}\right)\right], and the flux at the electrode surface (x=0) is J = -D \left(\frac{\partial C}{\partial x}\right)_{x=0} = \frac{C \sqrt{D}}{\sqrt{\pi t}}. The current is then i = n F A J, yielding the Cottrell equation. This model assumes one-dimensional diffusion to an infinite planar electrode with no convection or migration effects.^[34]^[35] For steady-state amperometry, where the current reaches a constant value, models account for enhanced mass transport via geometry or convection. At microelectrodes, hemispherical diffusion dominates, leading to the steady-state current

i_{ss} = 4 n F D C r,

where r is the electrode radius; this arises from solving the Laplace equation \nabla^2 C = 0 in spherical coordinates with boundary conditions C(r,t) = 0 and C(\infty,t) = C, resulting in a linear concentration gradient and constant flux J = D C / r.^[34] In convective systems, such as with a rotating disk electrode, the Levich equation quantifies the steady-state limiting current under laminar flow:

i_L = 0.620 n F A D^{2/3} \omega^{1/2} \nu^{-1/6} C,

where \omega is the rotation speed and \nu is the kinematic viscosity. This is derived by solving the convective-diffusion equation \frac{\partial C}{\partial z} = D \nabla^2 C with the Levich transformation for the Nernst diffusion layer thickness \delta \approx 1.61 D^{1/3} \omega^{-1/2} \nu^{1/6}, assuming a thin layer where convection is negligible and diffusion perpendicular to the surface dominates.^[34]^[36] The applied potential in amperometry is selected based on the Nernst equation, which relates the electrode potential to the surface concentrations of oxidized and reduced species:

E = E^0 + \frac{RT}{nF} \ln \left( \frac{[\ce{ox}]}{[\ce{red}]} \right),

where E^0 is the standard potential, R is the gas constant, and T is the temperature. For diffusion-controlled amperometry, a potential sufficiently beyond E^0 (typically E > E^0 + 0.1 V) ensures [\ce{ox}] \approx 0 at the surface, maximizing the current while minimizing kinetic limitations.^[34]^[37] These models rely on key assumptions, including semi-infinite diffusion (valid for short times or large solution volumes where the diffusion layer \sqrt{\pi D t} does not reach boundaries) and negligible ohmic drop (requiring sufficient ionic strength to minimize iR potential losses). Deviations occur in thin-layer cells or low-conductivity media, where finite diffusion or uncompensated resistance distorts the predicted currents.^[34]^[38]

Detection Techniques

Constant-Potential Amperometry

Constant-potential amperometry, also known as chronoamperometry, is an electrochemical technique where a constant potential is applied to the working electrode, and the resulting faradaic current is monitored over time to quantify electroactive species based on their diffusion-limited response.^[7] This method relies on a three-electrode setup consisting of a working electrode (typically platinum, gold, or carbon-based), a reference electrode (such as Ag/AgCl), and a counter electrode to maintain the applied potential and complete the circuit.^[39] The current initially decays following the potential step due to the establishment of a diffusion layer at the electrode surface, following the Cottrell equation in diffusion-controlled processes in unstirred solutions. Quantitative analysis typically uses the transient current at a fixed time after the step or the integrated charge, with steady-state currents achievable under convective mass transport conditions or using microelectrodes.^[33] The waveform employed is a simple potential step, transitioning from the initial open-circuit potential to the fixed working potential, with experiment durations commonly spanning seconds to minutes to capture the transient or steady-state current behavior.^[40] This configuration allows for real-time monitoring in batch analyses, particularly with macroelectrodes immersed in unstirred solutions, where natural diffusion governs mass transport without convective interference.^[41] For instance, in oxygen detection, Clark-type macroelectrodes apply a constant cathodic potential (around -0.6 V vs. Ag/AgCl) to reduce oxygen to hydrogen peroxide, enabling dissolved oxygen quantification in environmental or physiological samples.^[39] Similarly, glucose biosensors use macroelectrodes modified with glucose oxidase to generate hydrogen peroxide, which is then oxidized at a constant anodic potential (typically 0.5–0.7 V vs. Ag/AgCl), facilitating blood glucose monitoring in clinical settings.^[42] Selecting the appropriate potential is a critical practical step, often determined through preliminary cyclic voltammetry to identify the plateau region where the target analyte undergoes selective redox reaction without overlapping with interferents.^[43] Interference mitigation further enhances selectivity, achieved via electrode surface modifications such as self-assembled monolayers or nanomaterial coatings (e.g., carbon nanotubes) that promote analyte-specific interactions while repelling common interferents like ascorbate or urate.^[39] Additionally, selective permselective membranes, such as Nafion, can be applied to exclude anionic interferents, ensuring reliable performance in complex matrices like biological fluids.^[44]

Pulsed Amperometric Detection

Pulsed amperometric detection (PAD) is an advanced variant of amperometry that employs a time-dependent potential waveform to measure faradaic currents from electroactive species, particularly those that are challenging to detect under constant potential due to electrode fouling. Developed in the early 1980s by Dennis C. Johnson, PAD was initially introduced for the sensitive detection of simple alcohols at platinum electrodes in flow systems, addressing limitations of steady-state methods by incorporating periodic cleaning and reactivation steps to maintain electrode performance.^[45]^[46] Subsequent refinements by Johnson and William R. LaCourse in the late 1980s and 1990s extended PAD to non-electroactive analytes like carbohydrates, enabling their direct electrocatalytic oxidation in alkaline media without derivatization.^[47] This technique gained prominence in high-performance anion-exchange chromatography (HPAEC) coupled with PAD (HPAEC-PAD), revolutionizing the analysis of underivatized sugars and polyols. The core of PAD lies in its multi-step potential waveform, typically applied to noble metal electrodes such as gold or platinum, which facilitates in-situ oxide layer formation and removal to renew the electrode surface continuously. A standard three-step waveform for carbohydrate detection on a gold electrode includes a detection pulse at approximately +0.05 to +0.6 V (vs. Ag/AgCl) for analyte oxidation and current measurement (lasting 300–500 ms), followed by an oxidation pulse at +0.75 to +1.0 V (50–200 ms) to clean the electrode by forming a metal oxide layer that desorbs adsorbed residues, and a reduction pulse at -0.15 to +0.0 V (20–50 ms) to remove the oxide and restore catalytic activity.^[47]^[48] More advanced four-step waveforms incorporate an additional adsorption or activation step to enhance reproducibility, particularly for complex samples, with total cycle times of 100–500 ms to match chromatographic flow rates.^[21] These sequences prevent irreversible poisoning from oxidation products, contrasting with constant-potential amperometry by providing dynamic surface renewal for sustained sensitivity. PAD offers significant advantages for detecting polar, non-electroactive compounds such as carbohydrates, aliphatic alcohols, and amines, which exhibit weak responses under static potentials due to poor adsorption or rapid electrode deactivation. On noble metal surfaces in alkaline conditions, the pulsed waveform promotes electrocatalytic oxidation via transient metal hydroxide/oxide intermediates, enabling picomolar detection limits and high selectivity without interference from common mobile phase components.^[47]^[49] For instance, PAD facilitates the quantification of underivatized glucose and oligosaccharides in biological matrices by leveraging the reversible oxide chemistry on gold, yielding signal-to-noise ratios superior to UV detection.^[48] In practice, PAD instrumentation is commonly integrated as modular detectors in high-performance liquid chromatography (HPLC) systems, featuring waveform generators, potentiostats, and disposable or regenerable gold working electrodes paired with Ag/AgCl or Pd/H reference electrodes.^[21] Pulse timings are programmable via software like Chromeleon, allowing optimization for specific analytes (e.g., 400 ms detection for carbohydrates), and systems often include post-column pH adjustment or degassing to minimize baseline drift from dissolved oxygen.^[48] This setup ensures robust, automated operation for routine analyses, with electrode lifetimes extended to thousands of injections through the self-cleaning mechanism.

Specialized Variants

Biamperometry, also known as bipotentiometry, employs two identical working electrodes maintained at fixed potentials with a small applied voltage difference, typically 10-100 mV, to measure the current arising from the redox reaction between the analyte and titrant.^[50] The endpoint in titrations is determined by monitoring changes in the current, often reaching a minimum or zero value when the equivalence point is achieved, or by analyzing the ratio of oxidation to reduction currents for species like Fe(III)/Fe(II) during the titration process.^[51] This technique enhances selectivity in complex matrices by minimizing interference from non-redox active species, as the low potential difference avoids electrolysis of water or other background reactions.^[50] Hydrodynamic amperometry incorporates controlled convection to enhance mass transport to the electrode surface, primarily through rotating disk electrodes (RDEs) or flow cells in systems like flow injection analysis (FIA) or high-performance liquid chromatography (HPLC).^[52] In RDE setups, rotation generates a laminar flow pattern that establishes a steady-state diffusion layer thickness, governed by the Levich equation, allowing reproducible current responses proportional to analyte concentration under convective conditions.^[36] Flow cells, often integrated with amperometric detectors, facilitate rapid sample delivery and minimize dispersion, improving detection limits for electroactive species in continuous monitoring applications.^[52] These configurations are particularly advantageous for analytes with slow diffusion kinetics, as convection reduces the diffusion layer thickness compared to quiescent solutions.^[36] Microfluidic and nanoamperometry utilize ultramicroelectrodes (UMEs), with dimensions on the micrometer to nanometer scale, to achieve high spatial resolution and rapid temporal response for single-cell analysis.^[53] These electrodes enable amperometric detection of vesicular exocytosis events, such as neurotransmitter release from individual cells, with response times below 1 ms due to their small capacitance and minimal ohmic drop.^[54] In microfluidic devices, UMEs are integrated into channels for precise positioning near cell membranes, allowing real-time monitoring of quantal release without perturbing cellular function.^[55] This approach has been applied to study secretion dynamics in neuronal and endocrine cells, providing insights into vesicle fusion kinetics at the single-event level.^[53]

Applications

Analytical Chemistry

Amperometry serves as a key technique in analytical chemistry for the quantitative determination of electroactive species, leveraging the proportionality between measured current and analyte concentration to enable precise measurements in various matrices. This method is particularly valued for its sensitivity and selectivity in trace-level analysis, allowing for the detection of analytes at concentrations relevant to environmental and pharmaceutical monitoring. Calibration in amperometric methods typically involves constructing linear plots of steady-state current against analyte concentration under diffusion-controlled conditions, where the current is directly proportional to the diffusion-limited flux of the electroactive species to the electrode surface. These calibration curves facilitate accurate quantification, with limits of detection often reaching approximately 10^{-6} M for many species, such as glucose or heavy metals, depending on the electrode material and detection setup.^[56]^[57] Sample preparation for amperometric analysis is generally minimal in direct methods, requiring only the addition of a supporting electrolyte to ensure ionic strength and conductivity, which simplifies workflow and reduces contamination risks. However, for trace metal analysis, preconcentration techniques such as electrodeposition or complexation are employed to enhance sensitivity, while matrix effects from interferents like organic matter or other ions can necessitate dilution or masking agents to mitigate signal interference.^[56] Representative examples include the amperometric determination of heavy metals like Pb^{2+} in environmental water samples, where biosensor systems achieve detection limits as low as 9.38 \times 10^{-4} \mu g L^{-1} through enzyme inhibition mechanisms, enabling reliable assessment of contamination levels. Similarly, pharmaceuticals such as guaifenesin can be quantified in water matrices using flow injection amperometry with boron-doped diamond electrodes, offering limits of detection around 86 nmol L^{-1} and recoveries exceeding 94% without extensive pretreatment.^[58]^[59] Hyphenation with liquid chromatography (LC) enhances amperometry's utility for speciation analysis, where amperometric detection at the column outlet allows separation and selective quantification of species like inorganic and organic mercury, providing improved resolution for complex samples compared to standalone techniques.^[60]

Biosensing and Monitoring

Amperometric biosensors, particularly those employing enzymes such as glucose oxidase, have become pivotal in biological monitoring by detecting analytes through electron transfer processes at electrode surfaces. In these devices, the enzyme catalyzes the oxidation of the target substrate, generating a measurable current proportional to the analyte concentration; for instance, glucose oxidase immobilizes on the electrode to oxidize glucose, producing hydrogen peroxide that is electrochemically detected at a fixed potential.^[61] To enhance efficiency and reduce oxygen dependence, redox mediators like ferrocene derivatives shuttle electrons between the enzyme's active site and the electrode, enabling sensitive detection in physiological environments.^[62] This approach underpins commercial glucose biosensors for diabetic management, offering rapid response times and high selectivity for point-of-care testing.^[63] Implantable amperometric devices extend these principles to continuous in vivo monitoring, with the Clark electrode serving as a foundational example for oxygen sensing since its development in the 1950s. The Clark electrode operates by reducing oxygen at a platinum cathode covered by a semipermeable membrane, yielding a diffusion-limited current that correlates with partial oxygen pressure in tissues or blood.^[64] Building on this, continuous glucose monitors (CGMs) emerged in the late 1990s, with the first FDA-approved system in 1999 using enzyme-based amperometric sensors inserted subcutaneously to track interstitial glucose levels over several days.^[65] These devices, such as those from Medtronic and Dexcom, integrate oxidase enzymes with miniaturized electrodes, providing real-time data transmission to external receivers for glycemic control in diabetes patients.^[66] In environmental monitoring, portable amperometric sensors facilitate on-site detection of pollutants like nitrates in water bodies, employing nitrate reductase enzymes to biocatalyze nitrate reduction and generate quantifiable currents. These biosensors achieve detection limits below 1 mg/L, suitable for assessing eutrophication risks in rivers and groundwater, with deployments lasting up to two weeks in aquatic settings.^[67] For air quality, amperometric NOx sensors utilize electrochemical oxidation or reduction at solid-state electrodes to measure nitrogen oxides at parts-per-billion levels, enabling low-cost, compact monitoring in urban environments to track emissions from vehicles and industry.^[68] Such sensors differentiate NO2 from interferents like ozone through selective membrane coatings, supporting regulatory compliance and public health alerts.^[69] Wearable amperometric technologies have advanced by 2025 to noninvasively monitor sweat analytes during physical activity, focusing on lactate and electrolytes as indicators of metabolic stress and hydration status. Flexible electrochemical patches with enzyme-immobilized working electrodes detect lactate via lactate oxidase-mediated oxidation, correlating sweat levels (typically 5-20 mM during exercise) to blood equivalents for athlete performance tracking.^[70] These devices, often powered by flexible batteries, exemplify the shift toward personalized health monitoring in fitness applications.^[71]

Advantages and Limitations

Operational Benefits

Amperometry offers high sensitivity, enabling detection limits in the nanomolar (nM) range for various analytes, such as 50 nM for nitrite ions using specialized electrode configurations.^[72] This sensitivity arises from the direct measurement of faradaic currents proportional to analyte concentration, allowing quantification of low-abundance species without extensive preconcentration steps. Additionally, the technique provides rapid response times, often achieving real-time monitoring with transients resolving in milliseconds to less than one second, which facilitates dynamic studies of electrochemical processes.^[73]^[74] The operational simplicity of amperometry stems from its requirement for minimal sample preparation, typically involving only dilution or direct application to the electrode, which contrasts with more laborious methods like chromatography.^[75] Instrumentation is portable and cost-effective, relying on basic potentiostats that are less expensive than spectroscopic setups, making it suitable for field-deployable analyses.^[76] This ease of use supports high-throughput applications, with analytical frequencies exceeding 100 samples per hour in flow-based systems.^[75] Selectivity in amperometry is enhanced by precise control of the applied potential, which discriminates between analytes based on their redox potentials, minimizing interference from co-existing species.^[77] In biosensing contexts, enzyme immobilization on the electrode surface further improves specificity, as the biocatalyst selectively generates electroactive products from target substrates, enabling detection in complex matrices like biological fluids.^[78] Amperometry demonstrates versatility across diverse sample phases, including liquids through standard macroelectrodes and gases via solid electrolyte configurations for species like oxygen or nitrogen oxides.^[4] For solid samples, microelectrodes enable localized measurements, such as probing interfaces in battery materials or tissues, expanding applicability to heterogeneous systems.^[79]^[80]

Challenges and Considerations

One of the primary challenges in amperometry is electrode fouling, where proteins, polymers, or other biomolecules adsorb onto the electrode surface, blocking active sites and leading to signal decay and reduced sensitivity.^[81] This issue is particularly pronounced in biological matrices, such as during in vivo monitoring, where biofouling can cause impedance increases and hinder long-term sensor performance. Mitigation strategies include pulsed amperometric techniques that apply periodic cleaning potentials to oxidize or desorb foulants, thereby regenerating the surface and maintaining response stability.^[82] Additionally, the use of disposable electrodes, such as screen-printed or paper-based designs, avoids cumulative fouling by replacing the sensor after use, enhancing reliability in point-of-care applications.^[83] Interferences from matrix components represent another significant hurdle, as non-specific redox reactions of species like ascorbate, uric acid, or chloride can generate overlapping currents, compromising selectivity in complex samples.^[84] These interferences often arise in multi-component environments, such as physiological fluids, where unintended oxidations or reductions distort the target analyte signal.^[85] To address this, modified applied potentials can be employed to shift the detection window away from interfering species, minimizing their contribution while preserving analyte response.^[85] Selective membranes, including permselective or conductive polymer coatings, further enhance specificity by restricting access of interferents to the electrode surface.^[86] Reproducibility in amperometric measurements is often limited by variability in electrode history, including surface heterogeneity from prior use or fabrication inconsistencies, which can lead to inconsistent baseline currents and calibration slopes across devices.^[87] This electrode-to-electrode variation affects quantitative accuracy, particularly in batch-produced sensors. Preconditioning protocols, such as controlled potential cycling or short electrochemical pretreatments, standardize the surface state by removing oxides or adsorbed layers, thereby improving signal repeatability and long-term stability.^[88] For instance, constant potential preconditioning has been shown to prevent drift from sulfur adsorption in environmental sensors, ensuring consistent performance.^[89] Safety and scalability concerns arise from the use of high applied potentials, which risk unwanted electrolysis of water or solvent, producing hazardous byproducts like oxygen radicals or chlorine species that can degrade sensor materials or pose toxicity risks in biomedical contexts.^[90] In implantable devices, such reactions may also contribute to local inflammation or tissue damage. Miniaturization for implants introduces further challenges, including foreign body reactions that encapsulate the sensor, reducing analyte diffusion and signal fidelity over time.^[91] Balancing size reduction with sufficient sensitivity requires advanced fabrication techniques, such as nanochannel integration, to maintain performance while minimizing tissue disruption.^[92]

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Typically, the current is measured at a constant potential and this is referred to as amperometry. If a current is measured during controlled variations of the ...<|control11|><|separator|>
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An Introduction to Electrochemical Methods in Neuroscience - NCBI
The remainder of this chapter introduces the principles of amperometry and voltammetry, i.e., methods that involve the flow of current at electrodes under ...Missing: definition | Show results with:definition
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[PDF] Chapter 11
Amperometry and voltammetry, in which we measure current as a function of a fixed or variable potential, is the subject of Section 11D. Figure 11.5 ...
[12]
[PDF] Chapter 11
The focus of this chapter is on analytical techniques that use a measurement of potential, charge, or current to determine an analyte's concentration or to ...
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Ninety Years of Polarography - Heyrovský - Wiley Online Library
Feb 14, 2012 · Polarography came to existence first as “electrolysis with dropping mercury cathode” in 1922.[1,2] The current-potential curves measured ...
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Electrochemistry, Past and Present - ACS Publications
Titration at one indicator electrode was first reported by. Heyrovsky and Berezicky in 1929. A few years later, Kolthoff and his co-workers began their ...
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Amperometric Titration - an overview | ScienceDirect Topics
Amperometry was used for oxygen determination in the 1930s and 1940s. With the passage of time, much progress has been made in the amperometric method of ...Missing: early | Show results with:early
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History of blood gas analysis. IV. Leland Clark's oxygen electrode
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[PDF] The Potentiostat and the Voltage Clamp - The Electrochemical Society
In the early 1940s, Archie Hickling at the. University of Leicester, England, who was working in the field of electrochemistry, invented the potentiostat and ...Missing: amperometry | Show results with:amperometry
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Some Milestones in the 50-year History of Electrochemical Oxygen ...
In 1962 Clark and Lyons described the first biosensor. A Clark oxygen sensor was combined with glucose oxidase to detect glucose levels. In spite of the ...
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Electrochemical Glucose Sensors and Their Applications in ...
The first electrochemical glucose assay, based on reactions 1– 3, was described in 1961 by Malmstadt and Pardue. They added to reactions 1– 3 a fourth ...
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[PDF] Glucose-biosensors.pdf - Chemistry
Sci. 102, 29-45, 1962), Clark and. Lyons coined the term enzyme electrode. Clark's ideas became commercial reality in 1975 with the successful ...
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Pulsed Electrochemical Detection: Waveform Evolution
Using dc amperometry as a means of electrochemical (EC) detection following ... In this case, the origins of PED can be traced to a mutated dc ...
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High-performance anion-exchange chromatography with pulsed ...
Aug 9, 2025 · ... Hughes and Johnson (1981) first reported the use of PAD in platinum electrodes for detection of carbohydrates following liquid ...
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Anion Exchange Chromatography and Integrated Amperometric ...
During the 1980s and 1990s, electrochemical detection has gained acceptance as a method of choice for some important biomolecules.
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amperometric detection method (A00301) - IUPAC
A detection method in which the current is proportional to the concentration of the species generating the current. Source: PAC, 1990, 62, 2167.Missing: protocols nomenclature
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https://pmc.ncbi.nlm.nih.gov/articles/PMC4299072/
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Current Trends in Nanomaterial-Based Amperometric Biosensors
Possible functions that can be achieved with nanomaterial integration in amperometric biosensors. (1). Immobilization platform for biomolecules. (2).
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Development and recent advancement in microfluidics for point of ...
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Smartphone-enabled flow injection amperometric glucose ...
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Recent advances in the use of screen-printed electrodes in drug ...
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Measurement Uncertainty Estimation in Amperometric Sensors - NIH
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amperometry (09128) - IUPAC Gold Book
The current is usually faradaic and the applied potential is usually constant. Amperometry can be distinguished from voltammetry by the parameter being ...
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Chronoamperometry - an overview | ScienceDirect Topics
Chronoamperometry can therefore be used to measure current–time dependence for the diffusion controlled process occurring at an electrode. This varies with ...
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Chronoamperometry (CA) - Pine Research Instrumentation
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[PDF] ELECTROCHEMICAL METHODS
Apr 5, 2022 · We note here that the widely employed Clark oxygen electrode differs fundamentally from these devices (18, 63). The Clark device is similar ...
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Beyond steady-state conditions: Chronoamperometric state-of ...
Mar 15, 2024 · As common in electrochemistry literature, we refer to this technique as chronoamperometry throughout the study. ... Cottrell equation: [18], [19].
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Rotating Disk Electrode (RDE) Theory | Pine Research Instrumentation
Jan 27, 2025 · ... rotating disk electrode is given by the Levich equation: \displaystyle{i_{LC} = 0.620 n F A D_O^{2/. in terms of the concentration of the ...Missing: amperometry | Show results with:amperometry
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The Nernst Equation | Journal of Chemical Education
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Spatially limited diffusion coupled with ohmic potential drop and/or ...
In the present paper, we derive analytical approximations for the current-time behavior within the time interval corresponding to “semi-infinite diffusion”.
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https://www.basinc.com/manuals/EC_epsilon/Techniques/ChronoI/ca
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Chronoamperometry/chronocoulometry - BASi
In CA, the current is monitored as a function of time, whereas in CC, the charge is monitored as a function of time. It is important to note that the basic ...
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[PDF] Design and Development of Some Amperometric Biosensors
applying a constant potential of 0.65 V (vs. Ag/AgCl) in deaerated unstirred solutions for 20 min. The enzyme electrode was thoroughly rinsed with doubly ...
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Development of an Amperometric-Based Glucose Biosensor to ... - NIH
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Amperometry - an overview | ScienceDirect Topics
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(PDF) The use of Nafion to mitigate the “oxygen deficit” in glucose ...
... Constant potential amperometry (CPA) was used with a typical. three-electrode cell; all glucose detection was carried out at +0 V vs. SCE. De-aeration was ...
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https://doi.org/10.1016/0003-2670(81)90003-2
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Optimization of waveforms for pulsed amperometric detection of ...
Optimization of waveforms for pulsed amperometric detection of carbohydrates based on pulsed voltammetry. Click to copy article linkArticle link copied! William ...
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[PDF] Pulsed amperometric detection waveforms for carbohydrate ...
This waveform was first described in a publication by Rocklin et al ... Thermo Scientific Polishing amperometry cell gold working electrode https://tools.
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Pulsed amperometric detection of carbohydrates, amines and sulfur ...
A review is given of so-called pulsed amperometric detection at Au and Pt electrodes. Of greatest interest is the application of pulsed amperometric ...Missing: original | Show results with:original
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Theoretical and experimental study of the biamperometry for ...
It shows that small ΔE1/2 is favorable to construct the biamperometric detection system and to achieve high sensitivity and selectivity. Increasing ΔE leads to ...
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[PDF] Biamperometry in the Diffusion Current Region
As Fe(III) is formed, during the course of the titration, the current increases because the ratio. [Fe(III)]/[Fe(II)] increases. At the half-way titration point ...
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The hydrodynamics of the amperometric detector flow cell with a ...
Rotating the electrode generated a secondary fluid motion in the flow cell which manifested itself as vertical circulation of the solution present in the flow ...
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https://www.bioelectrochemical-soc.org/general/ultramicroelectrodes.php
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Recent developments concerning the investigation of exocytosis ...
First, ultramicroelectrodes and isolated single cells are globally the same sizes (∼μm). Second, ultramicroelectrodes have a fast time response (∼ms) and ...
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Single cell amperometry reveals curcuminoids modulate the release ...
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Measurement of Basal Neurotransmitter Levels Using Convolution ...
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Amperometric Method - an overview | ScienceDirect Topics
The device was designed using amperometric measurement and achieved high sensitivity and low detection limit (5 μmol L− 1). The results showed that the ...<|control11|><|separator|>
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Electrochemical Sensors | Analytical Chemistry - ACS Publications
To improve selectivity and stability, a differential amperometry protocol was adopted. Nitric oxide release rates of activated macrophages were subsequently ...
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Amperometric determination of cadmium, lead, and mercury metal ...
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Voltammetric and flow amperometric determination of drug guaifenesin in pharmaceutical and biological samples using screen-printed sensor with boron doped diamond electrode
### Summary of Amperometric Determination of Guaifenesin in Samples
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Liquid Chromatography with Pulsed Amperometric Detection for ...
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A Comprehensive Review of Glucose Biosensors Based on ... - NIH
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Recent advances in glucose monitoring utilizing oxidase ... - Frontiers
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Recent advances in amperometric glucose biosensors for in vivo ...
This review highlights recent developments in amperometric glucose biosensors for in vivo monitoring and also considers the remaining barriers which need to be ...
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Luminescent sensing and imaging of oxygen: Fierce competition to ...
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Continuous Glucose Monitoring Devices: Past, Present, and Future ...
In 1999, Minimed marketed the first CGM system that enabled recording of a patient's glucose values over a span of three days, which relied on repetitive sensor ...
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Introduction: History of Glucose Monitoring - NCBI - NIH
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Ion Selective Amperometric Biosensors for Environmental Analysis ...
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Amperometric Gas Sensors as a Low Cost Emerging Technology ...
Nov 22, 2017 · This review examines the use of amperometric electrochemical gas sensors for monitoring inorganic gases that affect urban air quality.
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Differentiating NO2 and O3 at Low Cost Air Quality Amperometric ...
This study deals with the differentiation and quantification of both nitrogen dioxide and ozone at ppb level when using amperometric gas sensors.
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Wearable and flexible electrochemical sensors for sweat analysis
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Diving into Sweat: Advances, Challenges, and Future Directions in ...
Aug 26, 2024 · This article introduces the rapid development of wearable and flexible sweat sensors, highlighting key milestones and various sensing strategies.
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(PDF) Advanced Wearable Devices for Monitoring Sweat ...
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Highly sensitive and selective amperometric determination of nitrite ...
Using amperometric method, we achieved the lowest detection of 50 nM nitrite for the first time. Further, the amperometric current increases linearly while ...
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Amperometry - an overview | ScienceDirect Topics
The electrochemical amperometry offers the capacity to distinguish ... The fundamental bases of the amperometric methods find their origins in the work ...
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Real-time amperometric measurements of zeptomole ... - PubMed
Amperometry with carbon-fiber microelectrodes provides a unique way to measure very small chemical concentration changes at the surface of biological cells.<|control11|><|separator|>
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Batch injection analysis with amperometric detection for ... - PubMed
Apr 22, 2021 · In addition, the minimum sample preparation (simple dilution) combined with a high analytical frequency (130 to 180 analyses per hour) can be ...
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Electrochemical Sensors - PMC - NIH
Electrochemical sensors are especially attractive owing to their simplicity, low cost, and portability compared to other detection strategies. Hrapovic et ...
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Immobilized Enzymes in Biosensor Applications - MDPI
Third-generation biosensors offer superior selectivity because they are able to operate in a potential range that is closer to the redox potential of the enzyme ...
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Addressing the Selectivity of Enzyme Biosensors - PMC - NIH
This review discusses the status of enzymatic biosensors in achieving accurate and selective measurements via direct biocatalytic and inhibition-based ...
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Principles, Development and Applications of Self-Referencing ...
Types of Amperometric Microelectrodes. The amperometric microelectrodes are solid-state, with a core of platinum, gold, or carbon as the electrolytic surface.
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https://pubmed.ncbi.nlm.nih.gov/9419871/
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Characterization of Electrode Fouling and Surface Regeneration for ...
This paper describes the characterization of electrode fouling and surface regeneration of an unmodified Pt metal electrode to accomplish sensitive and precise ...Missing: causes | Show results with:causes
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Improvements in amperometric detection of sulfite in food matrixes
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A green route for lignin-derived graphene electrodes: A disposable ...
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Interferences in a polypyrrole-based amperometric ammonia sensor
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An interference-free glucose biosensor based on a novel low ...
Another strategy to eliminate interferences from the mediated oxidation of glucose is to operate the glucose biosensor at a low enough applied potential so as ...
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A novel conductive membrane sensor protection technique to ...
In this paper, we present a novel conductive membrane encapsulation strategy designed to mitigate redox-active interferences while allowing redox-inactive ...
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Hitchhiker's Guide to Voltammetry: Acute and Chronic Electrodes for ...
Jan 27, 2017 · Therefore, the key to the success of this approach is to maintain reproducibility of electrode fabrication, a goal that may be more favorable ...
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The Effect of Preconditioning Strategies on the Adsorption of Model ...
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Planar carbon electrodes for real-time quantification of hydrogen ...
Electrodes were preconditioned using a constant potential amperometry methodology to prevent sensor drift resulting from elemental sulfur adsorption.
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Shattering the Water Window: Comprehensive Mapping of Faradaic ...
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Foreign Body Reaction to Implantable Biosensors: Effects of Tissue ...
Aug 25, 2015 · Sensor miniaturization in combination ... sensors was determined using both coated amperometric glucose sensors and microdialysis probes.
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Enhancing the Sensitivity of Needle-Implantable Electrochemical ...
Such performance proves that surface rebuilding of the WE can produce adequate sensitivity to afford device miniaturization as well as to bypass the need for ...