Improved porous silicon microarray based prostate specific antigen immunoassay by optimized surface density of the capture antibody

Enriching the surface density of immobilized capture antibodies enhances the detection signal of antibody sandwich microarrays. In this study, we improved the detection sensitivity of our previously developed P-Si (porous silicon) antibody microarray by optimizing concentrations of the capturing antibody. We investigated immunoassays using a P-Si microarray at three different capture antibody (PSA – prostate specific antigen) concentrations, analyzing the influence of the antibody density on the assay detection sensitivity. The LOD (limit of detection) for PSA was 2.5 ng mL⁻¹, 80 pg mL⁻¹, and 800 fg mL⁻¹ when arraying the PSA antibody, H117 at the concentration 15 μg mL⁻¹, 35 μg mL⁻¹, and 154 μg mL⁻¹, respectively. We further investigated PSA spiked into human female serum in the range of 800 fg mL⁻¹ to 500 ng mL⁻¹. The microarray showed a LOD of 800 fg mL⁻¹ and a dynamic range of 800 fg mL⁻¹ to 80 ng mL⁻¹ in serum spiked samples.     

Signal amplification strategy for sensitive immunoassay of prostate specific antigen (PSA) based on ferrocene incorporated polystyrene spheres

A new kind of signal amplification strategy based on ferrocene (Fc) incorporated polystyrene spheres (PS-Fc) was proposed. The synthesized PS-Fc displayed narrow size distribution and good stability. PS-Fc was applied as label to develop immunosensors for prostate specific antigen (PSA) after the typical sandwich immunoreaction by linking anti-PSA antibody (Ab₂) onto PS-Fc. After the fabrication of the immunosensor, tetrahydrofuran (THF) was dropped to dissolve PS and release the contained Fc for the following stripping voltammetric detection. PS-Fc as a new electrochemical label prevented the leakage of Fc and greatly amplified the immunosensor signal. In addition, the good biocompatibility of PS could maintain the bioactivity of the antibodies. The response current was linear to the logarithm of PSA concentration in the range from 0.01 ng mL⁻¹ to 20 ng mL⁻¹ with a detection limit of 1 pg mL⁻¹. The immunosensor results were validated through the detection of PSA in serum samples with satisfactory results.     

Aptamer-functionalized silver nanoparticles for scanometric detection of platelet-derived growth factor-BB

In this work, we reported a scanometric assay system based on the aptamer-functionalized silver nanoparticles (apt-AgNPs) for detection of platelet-derived growth factor-BB (PDGF-BB) protein. The aptamer and ssDNA were bound with silver nanoparticles by self-assembly of sulfhydryl group at 5′ end to form the apt-AgNPs probe. The apt-AgNPs probe can catalyze the reduction of metallic ions in color agent to generate metal deposition that can be captured both by human eyes and a flatbed scanner. Two different color agents, silver enhancer solution and color agent 1 (10 mM HAuCl₄ + 2 mM hydroquinone) were used to develop silver and gold shell on the surface of AgNPs separately. The results demonstrated that the formation of Ag core–Au shell structure had some advantages especially in the low concentrations. The apt-AgNPs probe coupled with color agent 1 showed remarkable superiority in both sensitivity and detection limit compared to the apt-AuNPs system. The apt-AgNPs system also produced a wider linear range from 1.56 ng mL⁻¹ to 100 ng mL⁻¹ for PDGF-BB with the detection limit lower than 1.56 ng mL⁻¹. The present strategy was applied to the determination of PDGF-BB in 10% serum, and the results showed that it had good specificity in complex biological media.     

Highly sensitive luminol electrochemiluminescence immunosensor based on ZnO nanoparticles and glucose oxidase decorated graphene for cancer biomarker detection

In this work, we reported a sandwiched luminol electrochemiluminescence (ECL) immunosensor using ZnO nanoparticles (ZnONPs) and glucose oxidase (GOD) decorated graphene as labels and in situ generated hydrogen peroxide as coreactant. In order to construct the base of the immunosensor, a hybrid architecture of Au nanoparticles and graphene by reduction of HAuCl₄ and graphene oxide (GO) with ascorbic acid was prepared. The resulted hybrid architecture modified electrode provided an excellent platform for immobilization of antibody with good bioactivity and stability. Then, ZnONPs and GOD functionalized graphene labeled secondary antibody was designed for fabricating a novel sandwiched ECL immunosensor. Enhanced sensitivity was obtained by in situ generating hydrogen peroxide with glucose oxidase and the catalysis of ZnONPs to the ECL reaction of luminol–H₂O₂ system. The as-prepared ECL immunosensor exhibited excellent analytical property for the detection of carcinoembryonic antigen (CEA) in the range from 10 pg mL⁻¹ to 80 ng mL⁻¹ and with a detection limit of 3.3 pg mL⁻¹ (S N⁻¹ = 3). The amplification strategy performed good promise for clinical application of screening of cancer biomarkers.     

Electrochemical stripping voltammetry of gold ions for development of ultra-sensitive immunoassay for chlorsulfuron

The paper reports a highly sensitive enzyme free electrochemical immunoassay (EFEIA) for the detection of herbicide chlorsulfuron. The assay is based upon oxidative gold nanoparticle (GNP) dissolution in an acidic solution. The consequent release of large amounts of gold (Au) metal ions after dissolution of gold nanoparticles tagged to antibody leads to the development of sensitive stripping voltammetry based immunoassay. The detection is made possible by the reduction of Au^3 + ions at the screen printed electrode surface followed by metal analysis by using the square wave voltammetry technique. The sensitivity of chlorsulfuron detection by competitive assay procedure was 6.7 pg mL^− 1 for EFEIA in marked contrast to optical detection using Standard ELISA procedure that gives a sensitivity of 4.97 ng mL^− 1.     

Simultaneous detection of four biomarkers with one sensing surface based on redox probe tagging strategy

In this paper, a simple and sensitive amperometric immunosensor for simultaneous detection of four biomarkers by using distinguishable redox-probes as signal tags was proposed for the first time. In sandwich immunoassay format, four kinds of capture antibodies (C-Ab) were immobilized by gold nanoparticles (AuNPs) electro-deposited on the surface of glass carbon electrode (GCE); four kinds of detection antibodies (D-Ab) labeled with different redox probes (including anthraquinone 2-carboxylic acid (Aq), thionine (Thi), ferrocenecarboxylic acid (Fc) and tris(2,2’-bipyridine-4,4’-dicarboxylic acid) cobalt(III) (Co(bpy)₃³⁺)), were combined with 3,4,9,10-perylenetetracarboxylic acid (PTCA), poly(diallyldimethylammonium chloride) (PDDA) and AuNPs functionalized carbon nanotubes, and served as signal tracer. When four target antigens were present, differential pulse voltammetry (DPV) scan exhibited four well-resolved peaks, each peak indicated one antigen, and its intensity was quantitative correlational to the concentration of corresponding analyte. To verify the strategy, four biomarkers for diagnosis of colorectal carcinoma, including carcinoembryonic antigen (CEA), carbohydrate antigen (CA) 19-9 CA125, and CA242, were used as model analytes, the immunosensor exhibited high selectivity and sensitivity, and peak current displayed good linear relationship to logarithm concentration in the ranges from 0.016 to 15 ng mL⁻¹ for CEA; 0.008 to 10 ng mL⁻¹ for CA19-9; 0.012 to 12 ng mL⁻¹ for CA125; 0.010 to 10 ng mL⁻¹ for CA242, and low detection limits of 4.2, 2.8, 3.3 and 3.8 pg mL⁻¹, respectively.     

One step electrochemically deposited nanocomposite film of chitosan-carbon nanotubes-gold nanoparticles for carcinoembryonic antigen immunosensor application

A simple and controllable one-step electrodeposition method for the preparation of a chitosan-carbon nanotubes-gold nanoparticles (CS-CNTs-GNPs) nanocomposite film was used to fabricate an immunosensor for detection of carcinoembryonic antigen (CEA). The porous three-dimensional CS-CNTs-GNPs nanocomposite film, which offered a large specific surface area for immobilization of antibodies, exhibited improved conductivity, high stability and good biocompatibility. The morphology of the formed nanocomposite film was investigated by scanning electron microscopy (SEM), and the electrochemical behaviors of the immunosensor were characterized by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). Under the optimal conditions, the proposed immunosensor could detect CEA in two linear ranges from 0.1 to 2.0 ng mL⁻¹ and from 2.0 to 200.0 ng mL⁻¹, with a detection limit of 0.04 ng mL⁻¹. The immunosensor based on CS-CNTs-GNPs nanocomposite film as the antibody immobilization matrix could exhibit good sensitivity, stability, and reproducibility for the determination of CEA.     

A sensitive impedance immunosensor based on functionalized gold nanoparticle-protein composite films for probing apolipoprotein A-I

A sensitive and label-free electrochemical impedance immunosensor via covalent coupling the antibody with functionalized gold nanoparticles (FAuNP) for probing apolipoprotein A-I was presented. The hybrid gold nanoparticles were prepared with a two-in-one strategy, i.e. via the stepwise employment of self-assembled monolayer (SAM) and sol-gel techniques, to improve the performance of such a label-free immunosensor, which was investigated by electrochemical impedance spectroscopy. It was found that this novel FAuNP immunosensor showed higher protein-loading capacity and better response properties (6-17 times) than that fabricated by normal SAM technique did. The remarkably improved properties of the immunosensor were ascribed to FAuNP with the larger surface-to-volume ratio, more free amino linkage groups, and the lower nonspecific protein adsorption. As a result, the thus-prepared antibody-modified immunosensor showed reproducible (R.S.D. = ±3.2%, n = 10) linear response to apolipoprotein A-I (Apo A-I) antigens in the range of 0.1-10 ng mL⁻¹. The detection limit of this immunosensor was 50 pg mL⁻¹ (corresponding to 1.8 pmol L⁻¹), which was two orders of magnitude lower than that of the traditional methods. These results exhibited the novel immunosensor had a high sensitivity, stability and selectivity for the determination of Apo A-I, especially in clinic microanalysis.     

Ultrasensitive electrochemical immunoassay for carcinoembryonic antigen based on three-dimensional macroporous gold nanoparticles/graphene composite platform and multienzyme functionalized nanoporous silver label

Three-dimensional macroporous gold nanoparticles/graphene composites (3D-AuNPs/GN) were synthesized through a simple two-step process, and were used to modify working electrode sensing platform, based on which a facile electrochemical immunoassay for sensitive detection of carcinoembryonic antigen (CEA) in human serum was developed. In the proposed 3D-AuNPs/GN, AuNPs were distributed not just on the surface, but also on the inside of graphene. And this distribution property increased the area of sensing surface, resulting in capturing more primary antibodies as well as improving the electronic transmission rate. In the presence of CEA, a sandwich-type immune composite was formed on the sensing platform, and the horseradish peroxidase-labeled anti-CEA antibody (HRP-Ab₂)/thionine/nanoporous silver (HRP-Ab₂/TH/NPS) signal label was captured. Under optimal conditions, the electrochemical immunosensor exhibited excellent analytical performance: the detection range of CEA is from 0.001 to 10 ng mL⁻¹ with low detection limit of 0.35 pg mL⁻¹ and low limit of quantitation (LOQ) of 0.85 pg mL⁻¹. The electrochemical immunosensor showed good precision, acceptable stability and reproducibility, and could be used for the detection of CEA in real samples. The proposed method provides a promising platform of clinical immunoassay for other biomolecules     

Development of an immunosensor for the detection of testosterone in bovine urine

In the presented work, a disposable immunosensor for the detection of testosterone, an endogenous steroid hormone, in bovine urine has been developed using screen-printed electrodes (SPEs). Due to concerns over the use of steroid hormones as growth promoters, the EU prohibits their use in food producing animals. Consequently, rigorous screening procedures have been implemented in all member states to detect the illegal administration of such compounds. Competitive immunoassays were developed, initially by enzyme linked immunosorbent assay (ELISA), and subsequently transferred to an electrochemical immunosensor format using disposable screen-printed carbon electrodes. Horseradish peroxidase (HRP) was the enzyme label of choice and chronoamperometric detection was carried out using a tetramethylbenzidine/hydrogen peroxide (TMB/H₂O₂) substrate system, at +100 mV. The EC₅₀ values obtained for the assay in buffer and urine gave relatively comparable results, 710 pg mL⁻¹ and 960 pg mL⁻¹, respectively. The linear range obtained for the assay in buffer extended from 0.03 ng mL⁻¹ to 40 ng mL⁻¹; while that in urine ranged from 0.03 ng mL⁻¹ to 1.6 ng mL⁻¹. The corresponding limits of detection (LOD) in buffer and urine were 26 pg mL⁻¹ and 1.8 pg mL⁻¹. Cross reactivity profiles of the antibody have been examined, with notable cross reactivities with 19-nortestosterone (11.6%) and boldenone (9.86%). Precision studies for the sensor demonstrated adequate reproducibility (CV < 13%, n = 3) and repeatability (CV < 9%, n = 3). Recovery data obtained showed good agreement between spiking studies and known concentrations of analyte. Sensors showed stability for 4 days at +4 °C. A sensitive, highly specific, inexpensive, disposable immunosensor, showing excellent overall performance for the detection of testosterone in bovine urine, has been developed.     

Determination of perfluorocarboxylic acids in water by ion-pair dispersive liquid–liquid microextraction and gas chromatography–tandem mass spectrometry with injection port derivatization

A novel technique for derivatization in a gas chromatograph injection port after a one-step extraction of trace perfluorocarboxylic acids (PFCAs) in water with ion pair formation during dispersive liquid–liquid microextraction (DLLME) was investigated. Tetrabutylammonium hydrogen sulfate (TBAHS) was used as the ion pair reagent. PFCA butyl ester derivatives were formed in the GC injection port and then analyzed using gas chromatography coupled to tandem mass spectrometry with negative chemical ionization. According to our analysis, the operative linear range for PFCA detection from 250 pg mL⁻¹ to 2 μg mL⁻¹ with a relative standard derivation (RSD) below 13%. Detection limits were achieved at the level of 37–51 pg mL⁻¹. This method was successfully applied for the analyzing of PFCAs in river water samples from urban and industrial areas without tedious pretreatment. The concentration range over which PFCAs were detected is from 0.6 ng mL⁻¹ to 604.9 ng mL⁻¹.     

Picogram-detection of estradiol at an electrochemical immunosensor with a gold nanoparticle|Protein G-(LC-SPDP)-scaffold

Low picograms of the hormone 17β-estradiol were detected at an electrochemical immunosensor. This immunosensor features a gold nanoparticle|Protein G-(LC-SPDP)¹-scaffold, to which a monoclonal anti-estradiol capture antibody was immobilised to facilitate a competitive immunoassay between sample 17β-estradiol and a horseradish peroxidase-labelled 17β-estradiol conjugate. Upon constructing this molecular architecture on a disposable gold electrode in a flow cell, amperometry was conducted to monitor the reduction current of benzoquinone produced from a catalytic reaction of horseradish peroxidase. This current was then quantitatively related to 17β-estradiol present in a sample. Calibration of immunosensors in blood serum samples spiked with 17β-estradiol yielded a linear response up to ∼1200 pg mL⁻¹, a sensitivity of 0.61 μA/pg mL⁻¹ and a detection limit of 6 pg mL⁻¹. We attribute these favourable characteristics of the immunosensors to the gold nanoparticle|Protein G-(LC-SPDP) scaffold, where the gold nanoparticles provided a large electrochemically active surface area that permits immobilisation of an enhanced quantity of all components of the molecular architecture, while the Protein G-(LC-SPDP) component aided in not only reducing steric hindrance when Protein G binds to the capture antibody, but also providing an orientation-controlled immobilisation of the capture antibody. Coupled with amperometric detection in a flow system, the immunosensor exhibited excellent reproducibility.     

Application of ZnO quantum dots dotted carbon nanotube for sensitive electrochemiluminescence immunoassay based on simply electrochemical reduced Pt/Au alloy and a disposable device

We report on a disposable microdevice suitable for sandwich-type electrochemiluminescence (ECL) detection of prostate specific antigen (PSA). The method is making use of ZnO quantum dots dotted carbon nanotube (ZnO@CNT) and simply electrochemical reduced Pt/Au alloy. The latter was selected as immunosensing probe to modify screen-printed carbon electrode, due to its excellent electrical property. For further ultrasensitive, low-potential and stable ECL detection, ZnO@CNT composite was first synthesized using a facile solvothermal method, and employed as signal amplification label. In this work, two working electrodes in one device were used for one determination to obtain more exact results based on screen-print technique. Taking advantage of dual-amplification effects of the Pt/Au and ZnO@CNT, this immunosensor could detect the PSA quantitatively, in the range of 0.001–500 ng mL⁻¹, with a low detection limit of 0.61 pg mL⁻¹. The resulting versatile immunosensor possesses high sensitivity, satisfactory reproducibility and regeneration. This simple and specific strategy has vast potential to be used in other biological assays.     

Using gold nanoparticles to improve the recovery and the limits of detection for the analysis of monohydroxy-polycyclic aromatic hydrocarbons in urine samples

We present a novel approach to improve the analytical figures of merit of solid-phase extraction high-performance liquid chromatography (SPE-HPLC) for the analysis of monohydroxy-polycyclic aromatic hydrocarbons in urine samples. The novel alternative substitutes the evaporation step that is currently used in SPE-HPLC methodology with a pre-concentration procedure that extracts metabolites with gold nanoparticles. The analytical potential of the new approach is evaluated with the following six metabolites: 9-hydroxyphenanthrene, 2-hydroxyfluorene, 1-hydroxypyrene, 6-hydroxychrysene, 3-hydroxybenzo[a]pyrene and 4-hydroxybenzo[a]pyrene. We demonstrate that the substitution of the evaporation step with the gold nanoparticles procedure improves the overall recoveries, the relative standard deviations of the average recoveries and the limits of detection of SPE-HPLC analysis. The overall recoveries of the studied metabolites varied from 59.7 ± 3.6% (2-hydroxyfluorene) to 92.3 ± 2.5% (6-hydroxychrysene). The relative standard deviations of the average recoveries were lower than 6%. The limits of detection were at the parts-per-trillion levels and varied from ∼2 pg mL⁻¹ (6-hydroxychrysene) to ∼18 pg mL⁻¹ (2-hydroxyfluorene).     

Electrochemical immunosensor based on nanoporpus gold loading thionine for carcinoembryonic antigen

Nanoporous gold (NPG) has recently received considerable attention in analytical electrochemistry because of its good conductivity and large specific surface area. A facile layer-by-layer assembly technique fabricated NPG was used to construct an electrochemical immunosensor for carcinoembryonic antigen (CEA). NPG was fabricated on glassy carbon (GC) electrode by alternatively assembling gold nanoparticles (AuNPs) and silver nanoparticles (AgNPs) using 1,4-benzenedimethanethiol as a cross-linker, and then AgNPs were dissolved with HNO₃. The thionine was absorbed into the NPG and then gold nanostructure was electrodeposited on the surface through the electrochemical reduction of gold chloride tetrahydrate (HAuCl₄). The anti-CEA was directly adsorbed on gold nanostructure fixed on the GC electrode. The linear range of the immunosensor was from 10 pg mL⁻¹ to 100 ng mL⁻¹ with a detection limit of 3 pg mL⁻¹ (S/N = 3). The proposed immunosensor has high sensitivity, wide linear range, low detection limit, and good selectivity. The present method could be widely applied to construct other immunosensors.     

An electrochemical immunosensor based on enzyme-encapsulated liposomes and biocatalytic metal deposition

A novel electrochemical immunosensor based on double signal amplification of enzyme-encapsulated liposomes and biocatalytic metal deposition was developed for the detection of human prostate specific antigen (PSA). Alkaline phosphatase (ALP)-encapsulated and detection antibody-functionalized liposomes were first prepared and used as the detection reagent. In the sandwich immunoassay, the model analyte PSA was first captured by anti-PSA capture antibody immobilized on the electrode and then sandwiched with the functionalized liposomes. The bound liposomes were then lysed with surfactant to release the encapsulated ALP, which served as secondary signal amplification means. ALP on the electrode surface initiated the hydrolysis of ascorbic acid 2-phosphate (AA-p) to produce ascorbic acid. The latter, in turn, reduced silver ions on the electrode surface, leading to deposition of the metal silver on the electrode surface. Linear sweep voltammetry (LSV) was chosen to detect the amount of the deposited silver. The results showed that the anodic stripping peak current was linearly dependent on the PSA concentration in the range of 0.01-100 ng mL⁻¹, and a detection limit as low as 0.007 ng mL⁻¹ can be obtained. Since the cut-off value of human PSA is 4 ng mL⁻¹, the proposed electrochemical immunosensor would be expected to gain widespread applications for the detection of PSA in clinical diagnosis.     

Resonance energy transfer between ZnCdHgSe quantum dots and gold nanorods enhancing photoelectrochemical immunosensing of prostate specific antigen

Gold nanorods (AuNRs) integrated with ZnCdHgSe near-infrared quantum dots (AuNRs-ZnCdHgSe QDs) were successfully synthesized and characterized by transmission electron microscope, X-ray photoelectron spectroscopy, and X-ray diffraction. A glassy carbon electrode was decorated with the aforementioned AuNRs-ZnCdHgSe QDs nanocomposite, which provides a biocompatible interface for the subsequent immobilization of prostate specific antibody (anti-PSA). After being successively treated with glutaraldehyde vapor and bovine serum albumin solution, a photoelectrochemical immunosensing platform based on anti-PSA/AuNRs-ZnCdHgSe QDs/GCE was established. The photocurrent response of ZnCdHgSe QDs was tremendously improved by AuNRs due to the effect of resonance energy transfer which can be deduced from the dependence of the enhanced efficiency on the AuNRs with different length-to-diameter ratios and spectral absorption characteristics. A maximum photocurrent was obtained when the absorption spectrum of AuNRs matched well with the emission spectrum of ZnCdHgSe QDs. A photoelectrochemical immunosensor for prostate specific antigen (PSA) was achieved by monitoring the photocurrent variation. The photocurrent variation before and after being interacted with PSA solution exhibits a good linear relationship with the logarithm of its concentration (logc_PSA) in the range from 1.0 pg mL⁻¹ to 50.0 ng mL⁻¹. The detection limit of this photoelectrochemical immunosensor is able to reach 0.1 pg mL⁻¹ (S/N = 3). Determining PSA in clinical human serum was also demonstrated by using the developed anti-PSA(BSA)/AuNRs-ZnCdHgSe QDs/GCE electrode. The results were comparable with those obtained from an enzyme-linked immunosorbent assay method.     

A novel immunoassay based on the dissociation of immunocomplex and fluorescence quenching by gold nanoparticles

This study reports a novel, simple and sensitive immunoassay using fluorescence quenching caused by gold nanoparticles coated with antibody. The method is based on a non-competitive heterogeneous immunoassay of human IgG conducted by the typical procedure of sandwich immunocomplex formation. Goat anti-human IgG was first adsorbed on polystyrene microwells, and human IgG analyte was captured by the primary antibody and then sandwiched by antibody labeled with gold nanoparticles. The sandwich-type immunocomplex was subsequently dissociated by the mixed solution of sodium hydroxide and trisodium citrate, the solution obtained, which contains gold nanoparticles coated with antibody, was used to quench fluorescence. The fluorescence intensity of fluorescein at 517 nm was inversely proportional to the logarithm of the concentration of human IgG in the dynamic range of 10-5000 ng mL⁻¹ with a detection limit of 4.7 ng mL⁻¹. The electrochemical experiments and the UV-vis measurements were applied to demonstrate whether the immunoglod was dissociated completely and whether the gold nanoparticles aggregated after being dissociated, respectively. The proposed system can be extended to detect target molecules such as other kinds of antigen and DNA strands, and has broad potential applications in disease diagnosis.     

Enzyme-catalyzed silver deposition on irregular-shaped gold nanoparticles for electrochemical immunoassay of alpha-fetoprotein

A new and disposable electrochemical immunosensor was designed for detection of alpha-fetoprotein (AFP), as a model analyte, with sensitivity enhancement based on enzyme-catalyzed silver deposition onto irregular-shaped gold nanoparticles (ISGNPs). The assay was carried out with a sandwich-type immunoassay protocol by using ISGNP-labeled anti-AFP antibodies conjugated with alkaline phosphatase (ALP–Ab₂) as detection antibodies. The enzymatically catalytic deposition of silver on the electrode could be measured by stripping analysis in KCl solution due to the Ag/AgCl solid-state voltammetric process. Several labeling protocols including spherical gold nanoparticle-labeled ALP–Ab₂ and ISGNP-labeled ALP–Ab₂ were investigated for determination of AFP, and improved analytical properties were achieved with the ISGNP labeling. With the ISGNP labeling method, the effects of incubation time and incubation temperature for antigen-antibody reaction, and deposition time of silver on the current responses of the electrochemical immunosensors were also monitored. Under optimal conditions, the electrochemical immunosensor exhibited a wide dynamic range from 0.01 ng mL⁻¹ to 200 ng mL⁻¹ with a detection limit of 5.0 pg mL⁻¹ AFP. The immunosensor displayed a good stability and acceptable reproducibility and accuracy. No significant differences at the 95% confidence level were encountered in the analysis of 10 clinical serum samples between the developed immunoassay and the commercially available electrochemiluminescent method for determination of AFP.     

A streptavidin functionalized graphene oxide/Au nanoparticles composite for the construction of sensitive chemiluminescent immunosensor

In this work, a novel streptavidin functionalized graphene oxide/Au nanoparticles (streptavidin/GO/AuNPs) composite is prepared and for the first time used to construct sensitive chemiluminescent immunosensor for the detection of tumor marker. The streptavidin/GO/AuNPs composite and the immunosensor are characterized using scanning electron microscopy, static water contact angle measurement and electrochemical impedance spectroscopy. The biofunctionalized composite has large reactive surface area and excellent biocompatibility, thus the capture antibody can be efficiently immobilized on its surface based on the highly selective recognition of streptavidin to biotinylated antibody. Using α-fetoprotein (AFP) as a model, the proposed chemiluminescent immunosensor shows a wide linear range from 0.001 to 0.1 ng mL⁻¹ with an extremely low detection limit down to 0.61 pg mL⁻¹. The resulting AFP immunosensor shows high detection sensitivity, fast assay speed, acceptable detection and fabrication reproducibility, good specificity and stability. The assay results of serum samples with the proposed method are in an acceptable agreement with the reference values. This work provides a promising biofunctionalized nanostructure for sensitive biosensing applications.