Amperometric Signal Research Articles

An electrochemical biosensor for mercury(II) detection based on DNA-Au bio-bar codes coupled with enzymatic dual signal amplification

Mercury ion (Hg2+) is one of the most toxic heavy metal pollutants, which not only causes enormous damage and pollution to environment, but also results in numerous irreversible impairments to the human health. Herein, a novel amperometric DNA biosensor was constructed for Hg2+ detection based on DNA-Au bio-bar codes coupled with enzymatic dual signal amplification. Carboxylated graphene oxide (GO − COOH) was selected as electrode modified material to provide anchoring sites for attachment DNA oligonucleotides. Substrate DNA was attached on the GO − COOH surface via covalent coupling interaction between amine (–NH2) groups on DNA and carboxyl (–COOH) groups on GO. AuNPs co-linked with bio-bar code DNA and target DNA via Au-S bonds were utilized as DNA-Au bio-bar codes, which labeled with HRP through avidin/biotin interaction. In the presence of object Hg2+, the target DNA on HRP-labeled DNA-Au bio-bar codes (HRP-DNA-Au bio-bar codes) hybridized with substrate DNA on GO − COOH via thymine-Hg2+-thymine (T-Hg2+-T) coordination chemistry. HRP-DNA-Au bio-bar codes catalyze hydroquinone oxidation to benzoquinone in presence of by hydrogen peroxide, generating an enhanced amperometric response signal. The current signal of biosensor is proportional to the logarithm of Hg2+ concentrations ranging from 25 pM to 10 μM with a detection limit of 10 pM (S/N = 3). Furthermore, without the need for target amplification, the prepared electrochemical DNA biosensor exhibited superior selectivity against other interfering metal ions, providing a facile and sensitive platform for Hg2+ detection in real environmental samples.

Comparison between Electropolymers of 3,5-Dihydroxybenzoic Acid and 2',6'-Dihydroxyacetophenone in Dimethyl Sulfoxide and Their Analytical Performance towards Selected Analytes with the Role of the Washing Liquid.

In the first part of this study, the electrochemical polymerization of two compounds, 3,5-dihydroxybenzoic acid and 2',6'-dihydroxyacetophenone, was compared in dimethyl sulfoxide solvent on platinum and glassy carbon electrodes. The voltammograms obtained showed remarkable differences between the two monomers and between the two electrode materials. The acetophenone derivative formed electropolymer remnants at the electrodes, while in the case of the benzoic acid derivative, practically no passivation occurred, and the scanning electron microscopic results reinforced this. A few stackings adsorbed only after electropolymerization from a highly concentrated solution of dihydroxybenzoic acid. As a modifying layer on the platinum and glassy carbon electrodes, the prepared films from 2',6'-dihydroxyacetophenone were tested for tributylamine in acetonitrile and in an aqueous solution of a redox-active compound, hydroquinone, during the stirring of the solution. More stable amperometric current signals could be reached with modified platinum than with glassy carbon, and the significant influence of the organic washing liquid after deposition was established via the study of noise level. In this respect, acetone was the best choice. The amperometric signals with the modified platinum obtained upon the addition of aliquots of the stock solution resulted in a 3.29 μM detection limit.

Quantification of lactic acid in wines using an amperometric biosensor

The objective of this study was to develop an analytical method that allows for the rapid and cost-effective detection and quantification of lactic acid in wines. The analysis of lactic acid is a time-consuming and costly process in the food industry. Therefore, the use of an enzymatic amperometric sensor that employs lactate oxidase (LacOx) immobilized on an oxygen electrode enabled the determination of lactic acid in wine. This study utilizes a voltage of −500mV and recorded the amperometric signal was obtained during oxygen consumption while lactic acid oxidation occurred in the sensor reactor. The detection time was at 10 s. A positive linear relationship was obtained between oxygen consumed as a function of time (mg O2/L x s−1). The lactic acid concentration ranged from 1 to 7 mM with a coefficient R2 = 0.994. The sensor exhibited good specificity KM = 0.675. The immobilized enzyme retained over 85% activity for 60 days, allowing for the reuse of the previously modified membrane up to 22 times. In comparison to other methods documented in the literature, the results demonstrate a favourable analytical quality. Furthermore, other aspects, such as the low residual formation, also exhibit a positive outcome. The analysis of lactic acid showed good correlation when compared to HPLC methodology, which occupies the same linear range. This validates the use of the sensor as an alternative method for quantifying lactic acid in wine. The value of this study lies in its presentation of a technique that is more straightforward to implement than traditional methods for quantifying lactic acid in grape wines.

MoS2 Modified Aluminum Electrode for Detection of Calcium Oxalate through Electrochemical Oxidation

The disposal of electronic waste (E-waste), encompassing discarded items such as computers, televisions, fridges, cellphones, and associated cabling, presents a significant challenge to both societal welfare and environmental health. Addressing e-waste management is critical to achieving several of the Sustainable Development Goals (SDGs). This study introduces a novel approach to repurposing aluminum from discarded electrical cables for the creation of high-efficiency electrodes that can detect oxalic acid in human urine samples. The process involves stripping 2mm thick cables to yield 7cm long aluminum rods. These rods are then coated with MoS2 nanoparticles via a hydrothermal synthesis technique. X-ray diffraction (XRD) analysis confirms the presence of MoS2 in the 1T phase, while Fourier-transform infrared spectroscopy (FTIR) and Raman spectroscopy provide insights into the vibrational modes and chemical composition. Field emission scanning electron microscopy (FESEM) imaging depicts the MoS2 as having a flower-like nanostructure composed of interwoven nanosheets. The resulting nanocomposite sensor demonstrates an enhanced electrical current response, along with notable selectivity and stability, likely due to the combined effects of the MoS2 materials. The sensor’s amperometric signal is directly correlated with the oxalate ion concentration within a range of 10 to 300 μM and exhibits a detection threshold as low as 2 μM. This sensor technique was evaluated alongside a standard colorimetric assay for measuring oxalic acid levels in urine specimens. Findings indicated a strong correlation (R=0.954) between oxalic acid concentrations assessed using the biosensor and those obtained through the colorimetric approach.Keywords: E-waste, Aluminum Electrodes, MoS2 Nanoparticles, Oxalic Acid Detection, and Electrochemical Sensing, Figure 1

Amperometric Signal Generation by Self-Doped Polyanilines for Ion-Selective Electrodes

Amperometric Signal Generation by Self-Doped Polyanilines for Ion-Selective Electrodes

Electrochemical biosensing platform based on AuNWs/rGO-CMC-PEDOT:PSS composite for the detection of superoxide anion released from living cells

Detection of superoxide anion (O2·-) levels holds significant importance for the diagnosis and even clinical treatments of oxidative stress-related diseases. Herein, we prepared a composite electrode material to encapsulate copper-zinc superoxide dismutase (SOD1) for biosensing of O2·-. The sensing material consists of gold nanowires (AuNWs), reduced graphene oxide (rGO), carboxymethyl cellulose (CMC) and PEDOT:PSS. CMC provides abundant -COOH to bind SOD1, with a high adsorption coverage of 1.499 × 10−9 mol cm−2 on the sensor surface. rGO and PEDOT endow the composite with significant conductivity, whereas PSS has antifouling capability. Moreover, AuNWs exhibit excellent electrical conductivity and a high aspect ratio, which promotes electron transfer, and ultimately enhances the catalytic performance of the enzyme. Meanwhile, SOD1(Cu2+) catalyzes the dismutation of O2·- to O2 and H2O2, and H2O2 is then electrochemically oxidized to generate amperometric signals for determination of O2·-. The sensor demonstrates outstanding detection performance for O2·- with a low detection limit of 2.52 nM, and two dynamic ranges (14.30 nM–1.34 μM and 1.34 μM–42.97 μM) with corresponding sensitivity of 0.479 and 0.052 μA μM−1cm−2, respectively. Additionally, the calculated apparent Michaelis constant (Kmapp) of 1.804 μM for SOD1 demonstrates the outstanding catalytic activity and the surface-immobilized enzyme's substrate affinity. Furthermore, the sensor shows the capability to dynamically detect the level of O2·- released from living HepG2 cells. This study provides an inovative design to obtain a biocompatible electrochemical sensing platform with plenty of immobilization sites for biomolecules, large surface area, high conductivity and flexibility.

Stability and reproducibility of the amperometric sensors for oxygen concentration analysis in the nitrogen gas mixtures

The paper presents experimentally obtained results on the long-term stability tests of the solid electrolyte amperometric sensor output signal during the operation in the O2 + N2 gas mixtures. These data prove the stability and reproducibility of the output signal when measuring the oxygen concentration in air for 8000 hours. The output signal variation during the tests did not exceed ± 2 %. We performed four heating / cooling cycles of different durations, which did not influence either sensor integrity or operation characteristics. The structure of the solid electrolyte sensor and the solid electrolyte/electrode interfacial layer remained unchanged during the tests. Dynamic characteristics of the sensor, including the response time, were stable.

Photoelectrochemical Enzyme Biosensor for Malate Using Quantum Dots on Indium Tin Oxide/Plastics as a Sensing Surface.

A photoelectrochemical biosensor for malate was developed using an indium tin oxide (ITO) layer deposited on a poly(ethylene terephthalate) plastic sheet as a transparent electrode material for the immobilization of malate dehydrogenase together with CdTe quantum dots. Different approaches were compared for the construction of the bioactive layer; the highest response was achieved by depositing malate dehydrogenase together with CdTe nanoparticles and covering it with a Nafion/water (1:1) mixture. The amperometric signal of this biosensor was recorded during irradiation with a near-UV LED in the flow-through mode. The limit of detection was 0.28 mmol/L, which is adequate for analyzing malic acid levels in drinks such as white wines and fruit juices. The results confirm that the cheap ITO layer deposited on the plastic sheet after cutting into rectangular electrodes allows for the economic production of photoelectrochemical (bio)sensors. The combination of NAD+-dependent malate dehydrogenase with quantum dots was also compatible with such an ITO surface.

High resolution electrochemical monitoring of small pH changes

Usually pH measurements are carried out using potentiometric technique. However, sensitivity of this method is low due to limitation set by Nernst equation – the moderate resolution is a significant problem in today demand for tracking small pH changes in various application areas.We propose a novel approach that allows electrochemical monitoring of pH changes with resolution reaching below 0.01 units in alkaline, acidic or close to neutral media. The proposed method uses all-solid state pH sensitive electrode with syringaldazine (Syr) modified multi-walled carbon nanotubes as the reference electrode modulating amperometric signal of Ag|AgCl electrode used as the working one. Due to high slope of the current vs. potential dependence for low-polarizable silver / silver chloride electrode relatively small changes of reference (pH-sensitive) electrode potential are transformed to pronounced changes of the recorded current. Ultimately, this results in significant increase in resolution of traced pH changes. Moreover, the current recorded is found to be linearly dependent on changes of H+ or OH− ions concentration in a broad pH range, which additionally contributes to increased resolution readout. Advantages of herein proposed approach were fully verified by monitoring minute changes of solution pH resulting from dissolution of carbon dioxide.

Bile acids modulate reinstatement of cocaine conditioned place preference and accumbal dopamine dynamics without compromising appetitive learning

Psychostimulants target the dopamine transporter (DAT) to elicit their psychomotor actions. Bile acids (BAs) can also bind to DAT and reduce behavioral responses to cocaine, suggesting a potential therapeutic application of BAs in psychostimulant use disorder. Here, we investigate the potential of BAs to decrease drug-primed reinstatement when administered during an abstinence phase. To do this, after successful development of cocaine-associated contextual place preference (cocaine CPP), cocaine administration was terminated, and animals treated with vehicle or obeticholic acid (OCA). When preference for the cocaine-associated context was extinguished, mice were challenged with a single priming dose of cocaine, and reinstatement of cocaine-associated contextual preference was measured. Animals treated with OCA demonstrate a significantly lower reinstatement for cocaine CPP. OCA also impairs the ability of cocaine to reduce the clearance rate of electrically stimulated dopamine release and diminishes the area under the curve (AUC) observed with amperometry. Furthermore, the AUC of the amperometric signal positively correlates with the reinstatement index. Using operant feeding devices, we demonstrate that OCA has no effect on contextual learning or motivation for natural rewards. These data highlight OCA as a potential therapeutic for cocaine use disorder.

RETRACTED: Multiple and simultaneous detection for cytokines based on the nanohole array by electrochemical sandwich immunoassay

Monitoring of cytokine storm markers is critical for determining the disease progression and treatment efficacy. In this study, we developed a novel fourplex electrochemical (EC) biosensor using nanohole array (NHA) structures containing several nanoholes on a single electrode for detecting cytokines with a low limit. A framework was developed for fabricating and optimizing multiplexed EC sandwich immunoassays using NHA electrodes because an NHA can enhance the EC detection of molecular binding by controlling the binding sites of captured biomolecules. For detecting multiple cytokines, the captured antibodies were immobilized on the surface-modified NHA and sandwich immunoassays with amperometric signal amplifications using poly-streptavidin horseradish peroxidase conjugates and 3,3′,5,5′-tetramethylbenzidine. Under the optimal conditions, the EC NHA immunosensors exhibited high sensitivity and specificity with a limit of detection of 1 pg/mL and a detection range of 1–300 pg/mL. The results revealed that the developed NHA sensor exhibits reliable EC characteristics and could be used to analyze multiple cytokines using small sample volume and subsequently developed for other applications.

Ultrasensitive Electrochemical Immunosensors Using Nanobodies as Biocompatible Sniffer Tools of Agricultural Contaminants and Human Disease Biomarkers.

Nanobodies (Nbs) are known as camelid single-domain fragments or variable heavy chain antibodies (VHH) that in vitro recognize the antigens (Ag) similar to full-size antibodies (Abs) and in vivo allow immunoreactions with biomolecule cavities inaccessible to conventional Abs. Currently, Nbs are widely used for clinical treatments due to their remarkably improved performance, ease of production, thermal robustness, superior physical and chemical properties. Interestingly, Nbs are also very promising bioreceptors for future rapid and portable immunoassays, compared to those using unstable full-size antibodies. For all these reasons, Nbs are excellent candidates in ecological risk assessments and advanced medicine, enabling the development of ultrasensitive biosensing platforms. In this review, immobilization strategies of Nbs on conductive supports for enhanced electrochemical immune detection of food contaminants (Fcont) and human biomarkers (Hbio) are discussed. In the case of Fcont, the direct competitive immunoassay detection using coating antigen solid surface is the most commonly used approach for efficient Nbs capture which was characterized with cyclic voltammetry (CV) and differential pulse voltammetry (DPV) when the signal decays for increasing concentrations of free antigen prepared in aqueous solutions. In contrast, for the Hbio investigations on thiolated gold electrodes, increases in amperometric and electrochemical impedance spectroscopy (EIS) signals were recorded, with increases in the antigen concentrations prepared in PBS or spiked real human samples.

Gold nanoparticles engineered nanoporous membrane with sandwich structure for sensitive electrochemical quantification of exosome

Exosome has been identified as an excellent potential biomarker for tumor diagnosis. Herein, electrochemical aptasensor for exosome detection was developed via a sandwich-structured nanoporous membrane which was fabricated by assembling multilayer gold nanoparticles (mulAu) and monolayer gold nanoparticles (monAu) on the opposite side of a nanoporous alumina membrane (NAM). The tumor-derived exosomes can be efficiently captured by the specific probes confined on the monAu side of the membrane. And the mulAu side that was tightly attached with a Si wafer allowed the electrochemical exosome detection by monitoring the amperometric signal of ferricyanide that transported across the nanochannel array of the membrane. After optimization of pore diameter of the NAM, base sequence of the capture probe and incubation temperature as well as incubation time for exosomes, this NAM-based electrochemical aptasensor was applied to the quantification of the MCF-7 cells-derived exosomes. The steady state current displayed linearity relationship with the concentration of the exosomes in a broad range of 103 to 107 particles μL−1, along with a detection limit of 2.8 × 102 particles μL−1. By changing capture probe, this sandwich-structured membrane could be further extended to label-free, sensitive and rapid quantification of other tumor-associated vesicles.

Nanomaterial-Doped Xerogels for Biosensing Measurements of Xanthine in Clinical and Industrial Applications.

First-generation amperometric xanthine (XAN) biosensors, assembled via layer-by-layer methodology and featuring xerogels doped with gold nanoparticles (Au-NPs), were the focus of this study and involved both fundamental exploration of the materials as well as demonstrated usage of the biosensor in both clinical (disease diagnosis) and industrial (meat freshness) applications. Voltammetry and amperometry were used to characterize and optimize the functional layers of the biosensor design including a xerogel with and without embedded xanthine oxidase enzyme (XOx) and an outer, semi-permeable blended polyurethane (PU) layer. Specifically, the porosity/hydrophobicity of xerogels formed from silane precursors and different compositions of PU were examined for their impact on the XAN biosensing mechanism. Doping the xerogel layer with different alkanethiol protected Au-NPs was demonstrated as an effective means for enhancing biosensor performance including improved sensitivity, linear range, and response time, as well as stabilizing XAN sensitivity and discrimination against common interferent species (selectivity) over time-all attributes matching or exceeding most other reported XAN sensors. Part of the study focuses on deconvoluting the amperometric signal generated by the biosensor and determining the contribution from all of the possible electroactive species involved in natural purine metabolism (e.g., uric acid, hypoxanthine) as an important part of designing XAN sensors (schemes amenable to miniaturization, portability, or low production cost). Effective XAN sensors remain relevant as potential tools for both early diagnosis of diseases as well as for industrial food monitoring.

A surprise from spatiotemporal analysis of exocytic events in chromaffin cells.

Exocytosis of individual secretory vesicles is mostly being studied by fluorescence imaging of vesicles loaded with a fluorescent marker, by amperometric detection of release of oxidizable transmitters such as catecholamines, or by measurements of membrane capacitance steps resulting from the increase in membrane area upon vesicle fusion. The relation between amperometric detection of catecholamine release and membrane capacitance changes has been characterized directly by patch amperometry and revealed that the amperometric pre spike foot signal (PSF) preceding the amperometric spike reflects release through a narrow fusion pore with fluctuations in catecholamine flux reflecting directly fluctuations of fusion pore conductance.

Metabolic pathway-based self-assembled Au@MXene liver microsome electrochemical biosensor for rapid screening of aflatoxin B1

Cytochrome P450 enzymes (CYPs) catalyze the production of aflatoxin B1 (AFB1) metabolites, which play an important role in carcinogenesis. In this study, we report a simple electrochemical liver-microsome-based biosensor using a composite of gold nanoparticles adsorbed on MXene (Au@MXene) for rapid screening of AFB1. Rat liver microsomes (RLMs) were directly adsorbed on the Au@MXene nanocomposite. The high conductivity, large specific surface area, and good biocompatibility of the Au@MXene nanocomposite enabled the direct electron transfer between the RLMs and the electrode and maintained the biological activity of the enzyme in the RLMs to a large extent. The metabolic behavior of the RLM biosensor that was developed for the electrocatalyst of AFB1 to its hydroxylation metabolite aflatoxin M1 (AFM1) was confirmed. Based on the change in the electrical signal generated by this metabolic behavior, we established the relationship between AFB1 content and amperometric (I-t) current signal. When the AFB1 concentration ranged from 0.01 μM to 50 μM, the AFB1 concentration was linearly related to the electrical signal with a limit of detection of 2.8 nM. The results of the recovery experiments for corn samples showed that the recovery and accuracy of the sensor were consistent with the UPLC-MS/MS method.

DNAzyme-driven bipedal DNA walker for label-free and signal-on electrochemical detection of amyloid-β oligomer

As a common technique for detecting AβO, the enzyme-linked immunosorbent assay (ELISA) method is time-consuming, high in cost, and poor in stability. Therefore, it is necessary to develop a highly sensitive, method-simple and low-cost method for the selective detection of AβO. Here, we created a novel signal-on and label-free electrochemical aptamer sensor for the detection of AβO based on a DNAzyme-driven DNA bipedal walking strategy. Compared with common DNA walkers, bipedal DNA walkers exhibit larger walking areas and faster walking kinetics, and provide higher amplification efficiency. The DNAwalker is powered by an Mg2+-dependent DNAzyme, and the binding-induced DNAwalker continuously clamps the MB, unlocking several active G-quadruplex-forming sequences. These G-quadruplexes can be further combined by hemin to generate a G-quadruplex/heme complex, resulting in an amperometric signal, resulting in a broad proportional band from 0.1 pM to 1 nM and an excellent detection range of 46 fM. A bipedal DNA walker aptamer sensor can detect human serum AβO with remarkable specificity, high reproducibility and practical application value.

Tracking a Major Egg Allergen to Assess Commercial Food Label Compliance: Towards a Simple and Fast Immunosensing Device.

An amperometric immunosensor was developed for the analysis of the major egg-white allergen ovotransferrin (Gal d 3) in commercial food products because the (accidental) intake, skin contact with, and/or inhalation of eggs can lead to severe disorders in allergic individuals. Employing a sandwich-type immunosensing strategy, screen-printed carbon electrodes (SPCE) were biomodified with anti-Gal d 3 (capture) antibodies, and the allergen's detection was achieved with anti-Gal d 3 antibodies labelled with horseradish peroxidase (HRP). The 3,3',5,5'-tetramethylbenzidine (TMB)/H2O2 reaction with HRP was used to obtain the electrochemical (amperometric) signal. An attractive assay time of 30 min and a remarkable analytical performance was achieved. The quantification range was established between 55 and 1000 ng·mL-1, with a limit of detection of 16 ng·mL-1. The developed method demonstrated good precision (Vx0 = 5.5%) and provided precise results (CV < 6%). The sensor also detected extremely low amounts (down to 0.010%) of egg. The analysis of seven raw and/or cooked egg and egg-white samples indicated that food processing influences the amount of allergen. Furthermore, to assure the compliance of product labelling with EU legislation, 25 commercial food ingredients/products were analysed. The accuracy of the results was confirmed through an ELISA assay. The stability of the ready-to-use sensing surface for 20 days allows a reduction of the reagents' volumes and cost.

Design of Nanostructured Hybrid Electrodes Based on a Liquid Crystalline Zn(II) Coordination Complex-Carbon Nanotubes Composition for the Specific Electrochemical Sensing of Uric Acid

A metallomesogen based on an Zn(II) coordination complex was employed as precursor to obtain a complex matrix nanoplatform for the fabrication of a high-performance electrochemical hybrid sensor. Three representative paste electrodes, which differ by the weight ratio between Zn(II) metallomesogen and carbon nanotubes (CNT), i.e., PE_01, PE_02 and PE_03, were obtained by mixing the materials in different amounts. The composition with the largest amount of CNT with respect to Zn complex, i.e., PE_03, gives the best electrochemical signal for uric acid detection by cyclic voltammetry in an alkaline medium. The amphiphilic structure of the Zn(II) coordination complex likely induces a regular separation between the metal centers favoring the redox system through their reduction, followed by stripping, and is characterized by enhanced electrocatalytic activity towards uric acid oxidation. The comparative detection of uric acid between the PE_03 paste electrode and the commercial zinc electrode demonstrated the superiority of the former, and its great potential for the development of advanced electrochemical detection of uric acid. Advanced electrochemical techniques, such as differential-pulsed voltammetry (DPV) and square-wave voltammetry (SWV), allowed for the highly sensitive detection of uric acid in aqueous alkaline solutions. In addition, a good and fast amperometric signal for uric acid detection was achieved by multiple-pulsed amperometry, which was validated by urine analysis.

High-Sensitivity Amperometric Dual Immunoassay Using Two Cascade Reactions with Signal Amplification of Redox Cycling in Nanoscale Gap.

Here, we report a high-sensitivity dual immunoassay using Lumulus amebocyte lysate (LAL) and blood coagulation cascade reactions with redox cycling in a nanoscale-gap electrode. Endotoxin and factor XIa were used as the label molecules for the immunoassay of two types of analytes to induce the LAL and coagulation cascade reactions, respectively, when each corresponding analyte existed in the sample solution. In addition to the signal amplification by the cascade reactions, we employed redox cycling in a nanoscale gap to achieve a highly sensitive assay. The nanoscale-gap electrode amplifies the amperometric signals from p-aminophenol liberated from artificial substrates in the final steps of the cascade reactions. First, the cross reaction between the LAL and coagulation cascade reactions was investigated. The results indicated that these cascade reactions did not efficiently proceed in a single solution owing to the cross reaction. Therefore, we selected to induce two cascade reactions in different solutions by bisecting the beads after the immunocomplex formation on the beads. The cross reactions of factor XIa with the LAL cascade reaction and of endotoxin with the coagulation cascade reaction were investigated. The effects of these cross reactions were revealed to be negligible by bisecting the beads before inducing the cascade reactions. Finally, a dual immunoassay for goat and human immunoglobulin G was performed, for which the limits of detection were 70 pg/mL (470 fmol/L) and 1.0 ng/mL (6.6 pmol/L), respectively. Thus, our dual immunoassay provides a sensitive platform for clinical diagnosis requiring detection of multiple analytes.