Disposable Immunosensor Research Articles

A disposable immunosensor for the detection of salivary MMP-8 as biomarker of periodontitis

This work describes the development of a novel voltammetric immunosensor for the detection of salivary MMP-8 at the point-of-care. The electrochemical platform was based on a graphene (GPH) screen-printed electrode (SPE) functionalized by gold-nanospheres (AuNSs) and antibodies against MMP-8 protein (anti-MMP-8). The functionalization with anti-MMP-8 was realized by using 11-mercaptoundecanoic acid (11-MUA), thanks to its ability to give strong sulfur bonds with its –SH end, and to cross-link the –NH2 groups of the antibody molecule with the other –COOH end, using the traditional EDC-NHS method. The voltammetric sensor showed good performances with a linear range of 2.5–300 ng mL−1, a LOD value of 1.0 ± 0.1 ng mL−1 and a sensitivity of 0.05 µA mL cm−2 ng−1. Moreover, the proposed immunosensor was tested in real saliva samples, showing comparable results to those obtained with the conventional ELISA method. The biosensor was single-use and cost-effective and required a small quantity of test medium and a short preparation time, representing a very attractive biosensor for MMP-8 detection in human saliva.

Point-of-care detection of Japanese encephalitis virus biomarker in clinical samples using a portable smartphone-enabled electrochemical "Sensit" device.

Japanese encephalitis (JE), a neglected tropical zoonotic disease prevalent in south-east Asian and western pacific countries, caused by the flavivirus JE virus (JEV), has a dearth of electrochemical point-of-care (PoC) diagnostic tools available to manage endemic breakouts. To overcome this, we have developed a screen-printed carbon electrode (SPCE) immunosensor for rapid PoC detection of JEV nonstructural 1 (NS1) antigen (Ag), found circulating in serum of infected individuals using a smartphone based portable "Sensit" device. The modification of SPCE surface with JEV NS1 antibody (Ab) was confirmed via observation of globular protein structures via scanning electron microscopy (SEM), increase in electrode surface hydrophilicity via contact angle measurement and decrease in current via differential pulse voltammetry (DPV). The fabrication and testing parameters were optimized based on highest current output obtained using DPV. The SPCE was tested for detection limit of target JEV NS1 Ag ranging from 1fM to 1μM, which was determined as 0.45 fM in spiked serum. The disposable immunosensor was also found to be highly specific in detecting JEV NS1 Ag over other flaviviral NS1 Ag. Finally, the modified SPCE was clinically validated by testing 62 clinical JEV samples using both a portable miniaturized electrochemical "Sensit" device coupled with a smartphone and a laboratory-based potentiostat. The results were corroborated with gold-standard RT-PCR and showed 96.77% accuracy, 96.15% sensitivity, and 97.22% specificity. Hence, this technique may further be developed into a one-step rapid diagnostic tool for JEV, especially in rural areas.

Using low-cost disposable immunosensor based on flexible PET screen-printed electrode modified with carbon black and gold nanoparticles for sensitive detection of SARS-CoV-2

To help meet the global demand for reliable and inexpensive COVID-19 testing and environmental analysis of SARS-CoV-2, the present work reports the development and application of a highly efficient disposable electrochemical immunosensor for the detection of SARS-CoV-2 in clinical and environmental matrices. The sensor developed is composed of a screen-printed electrode (SPE) array which was constructed using conductive carbon ink printed on polyethylene terephthalate (PET) substrate made from disposable soft drink bottles. The recognition site (Spike S1 Antibody (anti-SP Ab)) was covalently immobilized on the working electrode surface, which was effectively modified with carbon black (CB) and gold nanoparticles (AuNPs). The immunosensing material was subjected to a multi-technique characterization analysis using X-ray diffraction (XRD), transmission electron microscopy (TEM), and scanning electron microscopy (SEM) with elemental analysis via energy dispersive spectroscopy (EDS). The electrochemical characterization of the electrode surface and analytical measurements were performed using cyclic voltammetry (CV) and square-wave voltammetry (SWV). The immunosensor was easily applied for the conduct of rapid diagnoses or accurate quantitative environmental analyses by setting the incubation period to 10 min or 120 min. Under optimized conditions, the biosensor presented limits of detection (LODs) of 101 fg mL−1 and 46.2 fg mL−1 for 10 min and 120 min incubation periods, respectively; in addition, the sensor was successfully applied for SARS-CoV-2 detection and quantification in clinical and environmental samples. Considering the costs of all the raw materials required for manufacturing 200 units of the AuNP-CB/PET-SPE immunosensor, the production cost per unit is 0.29 USD.

Disposal Immunosensor for Sensitive Electrochemical Detection of Prostate-Specific Antigen Based on Amino-Rich Nanochannels Array-Modified Patterned Indium Tin Oxide Electrode.

Sensitive detection of prostate-specific antigens (PSA) in serum is essential for the prevention and early treatment of prostate cancer. Simple and disposable electrochemical immunosensors are highly desirable for screening and mobile detection of PSAs in high-risk populations. Here, an electrochemical immunosensor was constructed based on amino-rich nanochannels array-modified patterned, inexpensive, and disposable indium tin oxide (ITO) electrodes, which can be employed for the sensitive detection of PSA. Using an amino-group-containing precursor, a vertically ordered mesoporous silica nanochannel film (VMSF) containing amino groups (NH2-VMSF) was rapidly grown on ITO. When NH2-VMSF contained template surfactant micelle (SM), the outer surface of NH2-VMSF was directionally modified by aldehyde groups, which enabled further covalent immobilization of the recognitive antibody to prepare the immuno-recognitive interface. Owing to the charge-based selective permeability, NH2-VMSF can electrostatically adsorb negatively charged redox probes in solution (Fe(CN)63-/4-). The electrochemical detection of PSA is realized based on the mechanism that the antigen-antibody complex can reduce the diffusion of redox probes in solution to the underlying electrode, leading to the decrease in electrochemical signal. The constructed immunosensor can achieve sensitive detection of PSA in the range from 10 pg/mL to 1 μg/mL with a limit of detection (LOD) of 8.1 pg/mL. Sensitive detection of PSA in human serum was also achieved. The proposed disposable immunosensor based on cheap electrode and nanochannel array is expected to provide a new idea for developing a universal immunosensing platform for sensitive detection of tumor markers.

Ultrasensitive and Cost-Effective Detection of Neuropeptide-Y by a Disposable Immunosensor: A New Functionalization Route for Indium-Tin Oxide Surface.

Neuropeptide Y (NPY) is one of the most abundant neuropeptides in the human brain, and its levels in the blood change in neurodegenerative and neuroimmune disorders. This indicates that NPY may serve as a diagnostic and monitoring marker for associated disorders. In this paper, an electrochemical immunosensor was created to detect NPY biomarkers using a novel immobilization technique. The proposed biosensor system enables accurate, specific, cost-effective, and practical biomarker analysis. Indium tin oxide-coated polyethylene terephthalate (ITO-PET) sheets were treated with hexamethylene diisocyanate (HMDC) to covalently immobilize antibodies. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) techniques were used to analyze each step of the biosensors. The proposed NPY biosensor has a broad linear detection range (0.01-100 pg mL-1), a low limit of detection (LOD) (0.02968 pg mL-1), and a low limit of quantification (LOQ) (0.0989 pg mL-1). Atomic force microscopy (AFM) was used to support in the optimization process, study the surface morphology, and visualize it. Studies of repeatability, reproducibility, storage, and Kramers-Kronig transformation were conducted during electrochemical characterization. After analytical experiments, the biosensor's responses to human serum samples were evaluated. According to the obtained data, the error margin is small, and the created biosensor offers a great deal of promise for the clinical measurement of NPY.

Hierarchical Gold-Galinstan nanodendrites modified disposable immunosensor for the Label-free detection of KIM-1 by antibody immobilization on Staphylococcal protein a

In this work, we developed a novel Gold (Au)-Galinstan (Ga-In-Sn) nanodendrites modified label-free paper-based electrochemical immunosensor to detect one of the most prominent renal injury biomarkers known as Kidney injury molecule-1 (KIM-1). The Au-Galnstan dendritic nanostructure was obtained by a simple galvanic replacement method supported by polyethylene glycol (PEG) assisted co-reduction, where pH played an essential role in developing the dendritic morphology. Various physicochemical characterizations confirmed the successful formation of the nanodendrites in-depth. The large surface area, excellent synergistic charge transfer kinetics, and catalytic properties of the Au-Galinstan nanodendrites (Nds) modified paper electrode facilitated the oriented immobilization of the anti-KIM-1 antibodies involving Staphylococcal protein A (SPA). Under optimized conditions, the proposed immunosensor obtained an adequately low limit of detection (LOD) of 64 pg ml−1 in a wide linear range (0.1 ng ml−1 –1000 ng ml−1). The highly stable and easily reproducible sensor selectively identified KIM-1 in the presence of interferents. The high recovery rate obtained by the sensor in KIM-1 spiked human serum confirmed its applicability in a real clinical sample. Hence, the proposed immunosensor introduces an emerging platform to provide a simple, cost-effective, stable, and reliable detection of kidney injury-related markers.

Impedimetric Detection of Calreticulin by a Disposable Immunosensor Modified with a Single-Walled Carbon Nanotube-Conducting Polymer Nanocomposite.

A label-free impedimetric immunosensing system was constructed for ultrasensitive determination of the calreticulin (CALR) biological marker in human serum samples utilizing an electrochemical impedance spectroscopy analysis technique for the first time. The new biosensor fabrication procedure consisted of electrodeposition of single-walled carbon nanotubes (SWCNTs) incorporating polymerization of an oxiran-2-yl methyl 3-(1H-pyrrol-1-yl) propanoate monomer (Pepx) onto a low-cost and disposable indium tin oxide (ITO) electrode. The SWCNTs-PPepx nanocomposite layer was prepared onto the ITO after the one-step fabrication procedure. The fabrication procedure of the immunosensor and the characteristic biomolecular interactions between the anti-CALR and CALR were characterized by electrochemical analysis and morphological monitoring techniques. Under optimum conditions, the proposed biosensor was responsive to CALR concentrations over the detection ranges of 0.015-60 pg/mL linearly, and it had a very low detection limit (4.6 fg/mL) and a favorable sensitivity (0.43 kΩ pg-1 mL cm-2). The reliability of the biosensor system in clinical analysis was investigated by successful quantification of CALR levels in human serum. Moreover, the repeatability and reproducibility results of the biosensor were evaluated by using Dixon, Grubbs, T-test, and F-tests. Consequently, the proposed biosensor was a promising method for scientific, rapid, and successful analysis of CALR in human serum samples.

A Novel Disposable Immunosensor for Early Diagnosis of Cardiovascular Diseases

AbstractCardiovascular diseases are highly common cause of mortality and morbidity. It is envisaged that there is a relationship between cardiovascular diseases and ST2 (Suppression of Tumorigenicity 2) concentration of protein. In patients with cardiac insufficiency, the ST2 concentration was reported as higher than people without cardiac insufficiency. In this study ITO‐PET sheets (indium tin oxide covered terephthalate) was used as a working electrode for the fabrication of the immunosensor for ST2 protein. In the optimization experiments, important parameters responsible for biosensor fabrication have been optimized. After determining optimal conditions, reproducibility, repeatability, regeneration possibility, and analytical properties such as storage life are determined. The immunosensor showed a linear detection range for ST2 between 1–1500 fg/mL with a LOD value of 3.98 fg/mL. We also carried out SFI (single frequency impedance) experiments to monitor the specific interaction between ST2 and anti‐ST2, simultaneously. The developed immunosensor was successively applied to the determination of ST2 in commercial human serum sample.

A High Sensitive, Reproducible and Disposable Immunosensor for Analysis of SOX2

AbstractIn this study, an ITO (indium tin oxide) based biosensor was constructed to detect SOX2. SOX2 helps the regulation of cell pluripotency and is closely related to early embryonic development, neural and sexual differentiation. SOX2 is amplified and overexpressed in some malignant tumors such as squamous cell, lung, prostate, breast, esophageal cell, colon, ovarian, glioblastoma, pancreatic cancer, gastric cancer, head and neck squamous cell carcinoma. To generate a hydroxylated clean electrode surface, ITO electrodes were treated with NH4OH/H2O2/H2O. Later, ITO‐PET electrode surfaces were modified with 3‐glycidoxypropyl trimethoxysilane (3‐GOPS). Then, Anti‐SOX2 was covalently immobilized onto the electrode surfaces. 3‐GOPS concentration, Anti‐SOX2 concentration and incubation time, SOX2 incubation time were optimized. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were utilized in order to follow up the immobilization processes and the optimization steps of the biosensor. To characterize the analytical properties of constructed immunosensor; linear range, repeatability, reproducibility and regeneration studies were investigated. The linear range of the immunosensor was detected as 0.625 pg/mL–62.5 pg/mL. Square wave voltammetry technique was also applied to the biosensor. Storage life of the biosensor was determined for identifying the possible usability of the biosensor in clinical field. Finally, the designed biosensor was applied to the real human serum samples. The results obtained with the presented biosensor were also compared with ELISA results.

Impedimetric Immunosensor Utilizing Polyaniline/Gold Nanocomposite-Modified Screen-Printed Electrodes for Early Detection of Chronic Kidney Disease

The presence of small amounts of human serum albumin (HSA) in urine or microalbuminuria (30–300 µg/mL) is a valuable clinical biomarker for the early detection of chronic kidney disease (CKD). Herein, we report on the development of an inexpensive and disposable immunosensor for the sensitive, specific, and label-free detection of HSA using electrochemical impedance spectroscopy (EIS). We have utilized a simple one-step screen-printing protocol to fabricate the carbon-based three-electrode system on flexible plastic substrates. To enable efficient antibody immobilization and improved sensitivity, the carbon working electrode was sequentially modified with electropolymerized polyaniline (PANI) and electrodeposited gold nanocrystals (AuNCs). The PANI matrix serves as an interconnected nanostructured scaffold for homogeneous distribution of AuNCs and the resulting PANI/AuNCs nanocomposite synergically improved the immunosensor response. The PANI/AuNCs-modified working electrode surface was characterized using scanning electron microscopy (SEM) and the electrochemical response at each step was analyzed using EIS in a ferri/ferrocyanide redox probe solution. The normalized impedance variation during immunosensing increased linearly with HSA concentration in the range of 3–300 µg/mL and a highly repeatable response was observed for each concentration. Furthermore, the immunosensor displayed high specificity when tested using spiked sample solutions containing different concentrations of actin protein and J82 cell lysate (a complex fluid containing a multitude of interfering proteins). Consequently, these experimental results confirm the feasibility of the proposed immunosensor for early diagnosis and prognosis of CKD at the point of care.

Metal-enhanced chemiluminescence detection of C-reaction protein based on silver nanoparticle hybrid probes

A novel metal-enhanced chemiluminescence (MEC) signal tag was designed. By combining with a disposable immunosensor chip, an ultrasensitive immunoassay method was proposed for detection of C-reactive protein (CRP), an essential cerebrovascular disease marker. Two kinds of silver nanoparticle (AgNP) probes, DNA-hemin/DNA-A/biotin-DNA modified AgNPs (Probe A) and DNA-hemin/DNA-B modified AgNPs (Probe B) were prepared respectively. The MEC signal tag was formed by the link of Probe A and Probe B through the hybridization of DNA-A and DNA-B. The formed AgNP hybrid probes brought excellent CL signal amplification, due to the increased content of hemin molecules and the great MEC effect. The AgNP hybrid probes can be bound to the biotinylated antibody of sandwich immunocomplex for immunoassay of CRP. Under optimal conditions, the method showed a wide detection range of 7 × 10−7 to 0.07 mg mL−1 and a detection limit down to 0.05 ng mL−1. The results obtained with clinical serum samples were in acceptable agreement with reference values. The AgNP hybrid probes as well as the MEC-based immunoassay method showed great potentials in early clinical diagnosis of cerebrovascular disease.

Disposable Immunosensor Based on Electrochemiluminescence for Ultrasensitive Detection of Ketamine in Human Hair

An ultrasensitive disposable immunosensor based on a gold nanoparticles (AuNPs)/indium tin oxide (ITO) substrate has been constructed to detect ketamine (KET) conveniently. In this study, we obtained a AuNPs/ITO basic electrode by only a two-step ecofriendly drip-coating procedure. The self-assembled monolayers of AuNPs were used as a hetero-bifunctional cross-linker and activator, respectively. Ketamine antibody was used as a biorecognition determinant and immobilized onto the AuNPs/ITO through electrostatic interaction or Au–S or Au–N bonds. The architectures of AuNPs/ITO were characterized by transmission electron microscopy, scanning electron microscopy, cyclic voltammetry, and electrochemical impedance spectroscopy mainly. Electrochemiluminescence (ECL) was used to monitor the specific immune response between the antigen and antibody by measuring changes of intensity in the process of ECL. When the immune complex was incubated on the electrode, it blocked the electron communication and mass transfer,...

A carbon ink screen-printed immunoelectrode for Dengue virus NS1protein detection based on photosynthesized amine gold nanoparticles

Dengue virus (DENV) is a reemerging mosquito-borne disease that is endemic in more than 125 countries, affecting 200 million people per year. Screening testing has been a good attempt to minimize the impact caused by high morbity and mortality rates of DENV. In this study, a simple and disposable label-free electrochemical immunosensor based on a carbon ink graphite screen-printed electrode (SPE) one-step fabricated was developed for detection of non-structural 1 protein (NS1). The SPE surface was modified by drop casting, depositing a colloidal suspension containing amine-functionalized gold nanoparticles (AuNP-NH2). AuNPs were synthetized by a photoinduced physical method, illuminating preformed gold seeds with a light-emitting diode (LED,) at blue region, by using the polyethyleneimine (NH2) as reductor and stabilizing agent. UV-VIS spectroscopy and Transmission Electron Microscopy (TEM) were used to characterize the amine AuNPs. Electrocatalytic activity of AuNPs allowed more sensitivity for a label-free detection of NS1 by square wave voltammetry (SWV), with linear response from 0.1 to 2 µg mL-1. It was found a good linearity (coefficient of correlation of 0.995 (p<0.01) and a limit of detection of 0.03 µg mL-1 NS1 for analytical responses. AuNP-NH2 synthesis provided an easy oriented immobilization of anti-NS1 antibodies by Fc portion, resulting in a simple fabrication immunosensor with relative high performance and feasibility for early diagnostic of DENV.

Ultrasensitive environmental assessment of xeno-estrogens in water samples using label-free graphene immunosensors

There is a growing interest in the possible environmental health impact posed by endocrine-disrupting chemicals (EDCs). A challenge to the field of endocrine disruption is that these substances are diverse and may not appear to share any structural similarity other than usually being low molecular mass (<1000 Da) compounds. Here we demonstrate the effectiveness of sensor device for the detection of low molecular weight, poorly water soluble, estrogenic compounds E1, E2 and EE2, fabricated by electropolymerization over graphene screen printed electrode (SPE).The PANI/Gr-SPE-devices displayed linear responses to estrogenic substances, in EIS assays, from 0.0975 ng/L to 200 ng/L in water samples, with a detection limit of 0.043 pg/L for E1, 0.19 ng/L for E2 and 0.070 pg/L for EE2 which is lower than other current biosensing techniques. This portable, disposable immunosensor offers a solution for immediate measurement at sample collection sites, due to its excellent sensitivity and selectivity when testing water samples obtained directly from rivers and waste water treatment facilities. The simple screen printing production method will enable the low cost, high volume production required for this type of environmental analysis.

A disposable immunosensor using ITO based electrode modified by a star-shaped polymer for analysis of tumor suppressor protein p53 in human serum

Label-free immunosensor based on tetra armed star-shaped poly(glycidylmethacrylate) (StarPGMA) modified disposable ITO electrode was developed for detection of p53 protein, an important colorectal cancer biomarker. This disposable biosensor was fabricated by spin-coating technique using star-shaped StarPGMA with epoxy side groups. After formation of a stable film with epoxy ends, anti-p53 antibodies were bound to these groups covalently. Stepwise modification of the electrode was followed by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) studies. The electrochemical performance of the immunosensor was studied by EIS. Furthermore, the antibody and antigen coupling was monitored by single frequency impedance (SFI) technique. The immunosensor showed a low limit of detection (7 fg/mL) and a linear detection range between 0.02 pg/mL and 4 pg/mL. Additionally, atomic force microscopy (AFM) and scanning electron microscopy (SEM) were utilized for monitoring of immunosensor surface at different stages of fabrication. The antibody coupling on the electrode surface was proved by Fourier-transform infrared spectroscopy (FTIR) and Raman spectroscopy. Furthermore, the proposed immunosensor had good reproducibility and repeatability.

Label-free Detection of Prostate Specific Antigen at a Screen-printed Immunosensor Modified with a Nanostructured Gold Layer

The development of a disposable immunosensor based on a hybrid gold–carbon screen-printed electrode array is described. The electrodeposition of gold is shown to lead to a field of nanocrystals that facilitates antibody immobilization and enhances electron transfer. A label-free analysis protocol based on ferrocyanide has been developed for the detection of Prostate Specific Antigen in the fg mL−1 range.

Cost-Effective and Handmade Paper-Based Immunosensing Device for Electrochemical Detection of Influenza Virus.

Although many studies concerning the detection of influenza virus have been published, a paper-based, label-free electrochemical immunosensor has never been reported. Here, we present a cost-effective, handmade paper-based immunosensor for label-free electrochemical detection of influenza virus H1N1. This immunosensor was prepared by modifying paper with a spray of hydrophobic silica nanoparticles, and using stencil-printed electrodes. We used a glass vaporizer to spray the hydrophobic silica nanoparticles onto the paper, rendering it super-hydrophobic. The super-hydrophobicity, which is essential for this paper-based biosensor, was achieved via 30–40 spray coatings, corresponding to a 0.39–0.41 mg cm−2 coating of nanoparticles on the paper and yielding a water contact angle of 150° ± 1°. Stencil-printed carbon electrodes modified with single-walled carbon nanotubes and chitosan were employed to increase the sensitivity of the sensor, and the antibodies were immobilized via glutaraldehyde cross-linking. Differential pulse voltammetry was used to assess the sensitivity of the sensors at various virus concentrations, ranging from 10 to 104 PFU mL−1, and the selectivity was assessed against MS2 bacteriophages and the influenza B viruses. These immunosensors showed good linear behaviors, improved detection times (30 min), and selectivity for the H1N1 virus with a limit of detection of 113 PFU mL−1, which is sufficiently sensitive for rapid on-site diagnosis. The simple and inexpensive methodologies developed in this study have great potential to be used for the development of a low-cost and disposable immunosensor for detection of pathogenic microorganisms, especially in developing countries.

An electrochemical immunosensor for brain natriuretic peptide prepared with screen-printed carbon electrodes nanostructured with gold nanoparticles grafted through aryl diazonium salt chemistry

A sensitive amperometric immunosensor has been prepared by immobilization of capture antibodies onto gold nanoparticles (AuNPs) grafted on a screen-printed carbon electrode (SPCE) through aryl diazonium salt chemistry using 4-aminothiophenol (AuNPs-S-Phe-SPCE). The immunosensor was designed for the accurate determination of clinically relevant levels of B-type natriuretic peptide (BNP) in human serum samples. The nanostructured electrochemical platform resulted in an ordered layer of AuNPs onto SPCEs which combined the advantages of high conductivity and improved stability of immobilized biomolecules. The resulting disposable immunosensor used a sandwich type immunoassay involving a peroxidase-labeled detector antibody. The amperometric transduction was carried out at −0.20V (vs the Ag pseudo-reference electrode) upon the addition of hydroquinone (HQ) as electron transfer mediator and H2O2 as the enzyme substrate. The nanostructured immunosensors show a storage stability of at least 25 days, a linear range between 0.014 and 15ngmL−1, and a LOD of 4pgmL−1, which is 100 times lower than the established cut-off value for heart failure (HF) diagnosis. The performance of the immunosensor is advantageously compared with that provided with immunosensors prepared by grafting SPCE with p-phenylendiamine (H2N-Phe-SPCE) and attaching AuNPs by immersion into an AuNPs suspension or by electrochemical deposition, as well as with immunosensors constructed using commercial AuNPs-modified SPCEs. The developed immunosensor was applied to the successful analysis of human serum from heart failure (HF) patients upon just a 10-times dilution as sample treatment.

Disposable Electrochemical Immunosensor Based on Dual-Labeled Gold Nanoprobe (Ab-Au-HRP) for Detection of Alpha-Fetoprotein

A disposable electrochemical immunosensor for detection of human alpha-fetoprotein (AFP) has been developed in this study, using monoclonal antibody against AFP and horseradish peroxidase co-modified colloidal gold nanoparticles (Ab-Au-HRP) as trace label and screen-printed carbon electrodes (SPCEs) as signal transducers. The immunosensor was based on a sandwich immunoassay scheme in which the nanoprobe Ab-Au-HRP was dispensed on the polyester membrane as conjugate pad. The other monoclonal antibody against AFP, as capture antibody, was immobilized onto the nitrocellulose membrane which was mounted on the carbon surface of the screen-printed electrodes. An absorbent paper was mounted downstream to draw the sample to successively flow through the conjugate, capture and absorbent pads. The resulting sandwich immune reaction was evaluated by electrochemical detection, through addition of a single-component substrate for HRP. Under optimum conditions, the current value was linearly increased with an increased AFP concentration from 0.5 to 350 ng/mL, at a relatively low detection limit of 0.25 ng/mL at 3 sigma. Excellent analytical performance, fabrication reproducibility and operational stability for the proposed immunosensor indicated its promising application in clinical diagnostics and self-exams.

Construction of novel electrochemical immunosensor for detection of prostate specific antigen using ferrocene-PAMAM dendrimers

In this study, an immunosensor was designed to utilize for the detection of prostate specific antigen (PSA) based on three different generations (G1, G2 and G3) of ferrocene (Fc) cored polyamidiamine dendrimers (Fc-PAMAM) gold (Au) electrode. The self-assembled monolayer principle (SAM) was used to fabricate the sensitive, selective and disposable immunosensor electrodes. In electrode fabrication cysteamine (Cys) was the first agent covalently linked on the Au electrode surface. Immobilized redox center (ferrocene) cored PAMAM dendrimers served as a layer for the further binding of biological components. The monoclonal antibody of PSA (anti-PSA) was covalently immobilized on dendrimers which were attached onto the modified Au surface (Au/Cys/Fc-PAMAMs/anti-PSA). PSA levels were quantitatively analyzed by using electrochemical differential pulse voltammetry (DPV) whose lowest detection limit was calculated as 0.001ngmL−1. The Au/Cys/FcPAMAM/anti-PSA immunosensor showed excellent performance for PSA at the pulse amplitude; 50mV and the scan rate; 10mV/s in a wide linear concentration range of 0.01ng–100ngmL−1. Analytical performance and specificity assays were carried out using human serum and different proteins.