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Cortisol, a steroid hormone, has been confirmed as a kind of biomarker that reflects the stress response of psychobiology and related adverse health outcomes. Therefore, a sensitive and selective non-invasive assay to detect cortisol is needed. In this work, we developed a new photoelectrochemical immunosensor for determining cortisol levels in human saliva, based on evident photocurrent of the low-toxic CuInS2 quantum dots (CIS QDs) immobilized on the surface of TiO2 and significant signal amplification of the CdS/melamine network as a label. The photocurrent depended on the amount of label competing with cortisol antigen to bind the antibody on the electrode. The photocurrent decreased with the increment of cortisol concentration. Under the optimal conditions, this cortisol biosensor exhibited a broad linear range (0.01–100 ng/mL) with a low detection limit (7.8 pg/mL). Moreover, the methodology was implemented with good stability and high repeatability. [ABSTRACT FROM AUTHOR]

In this paper, a simple and feasible immunoassay protocol with signal amplification for sensitive determination of alpha fetoprotein (AFP, used as a model) by using new enzyme-induced Fenton reaction (EIFR) accompanying the oxidation of o-phenylenediamine (OPD) system. We discovered that glucose oxidase (GOx) has the ability to stimulate in situ Fenton reaction coupling the oxidation of OPD. GOx-catalyze of glucose results in the formation of hydrogen peroxide (H2O2). It can catalytically oxidize Fe2+to Fe3+, meanwhile producing hydroxyl radicals (•OH). The latter effectively initiated the catalytic oxidation of o-phenylenediamine (OPD) to 2,2-aminoazobenzene (DAP), resulting in the production of an electrochemical signal. On the basis of EIFR-OPD system, a new immunoassay protocol with GOx/anti-pAb2-conjugated gold nanoparticle (Ab2-AuNP-GOx) detection antibody can be designed for the detection of target AFP on capture antibody–functionalized transparent 96-well polystyrene microplate, monitored by recording the current peak of the generated DAP. The electrochemical current shows to be positively correlated with the concentration of target AFP. Under optimal conditions, the proposed assay exhibited good electrochemical responses for detecting target AFP in the range of 1.0 pg mL−1 to 100 ng mL−1 with a detection limit (LOD) of 0.5 pg mL−1 (0.5 ppt) estimated at the 3Sblank level. Additionally, the precision, reproducibility, specificity, and method accuracy were also investigated with acceptable results. Importantly, the EIFR-OPD system can be further extended for the determination of other protein or biomarkers by controlling the specific recognition system (antibody or aptamer). [ABSTRACT FROM AUTHOR]

We report on a protocol for a simultaneous competitive immunoassay for tetracycline (TC) and chloramphenicol (CAP) on the same sensing interface. Conjugates of TC and of CAP with bovine serum albumin were first co-immobilized on a glassy carbon electrode modified with gold nanoparticles. In parallel, monoclonal anti-TC and anti-CAP antibodies were conjugated onto CdS and PbS nanoclusters, respectively. In a typical assay, the immobilized haptens and the added target analytes competed for binding to the corresponding antibodies on the nanoclusters. Subsequently, Cd(II) and Pb(II) ions are released from the surface of the corresponding nanoclusters by treatment with acid and then were detected by square wave anodic stripping voltammetry. The currents at the peak potentials for Cd(II) and Pb(II) were used as the sensor signal for TC and CAP, respectively. This multiplex immunoassay enables the simultaneous determination of TC and CAP in a single run with dynamic ranges from 0.01 to 50 ng mL for both analytes. The detection limits for TC and for CAP are 7.5 pg mL and 5.4 pg mL, respectively. No obvious nonspecific adsorption and cross-reactivity was observed in a series of analyses. Intra-assay and inter-assay coefficients of variation were less than 10 %. The method was evaluated by analyzing TC and CAP in spiked samples of milk and honey. The recoveries range from 88 % to 107 % for TC, and from 91 % to 119 % for CAP. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

A novel, enzyme-free amperometric immunoassay of biomarkers with sensitive enhancement was designed by using gold nanoflower-labeled detection antibodies toward the catalytic reduction of p-nitrophenol and redox cycling of p-aminophenol on a graphene-based Au(111) platform. [ABSTRACT FROM AUTHOR]

[Display omitted] • Colorimetric and photothermal dual-signal immunoassay biosensor was developed. • Affordable detection of AFB 1 was achieved by a common thermometer-based photothermal method. • Application of immunoassay in food samples with high sensitivity. In this work, a split-type dual-mode (colorimetric/photothermal) immunoassay method was designed for point-of-care testing (POCT) detection of mycotoxins (aflatoxin B 1 , AFB 1 as the model analyte) in foodstuffs based on Pt supported on nitrogen-doped carbon amorphous (Pt-CN). The as-synthesized Pt-CN exhibits excellent peroxidase-mimicking activity, which can catalyze the oxidization of 3,3′,5,5′-tetramethylbenzidine (TMB) into TMB ox with sensitive colorimetric readout in the presence of hydrogen peroxide (H 2 O 2). Moreover, the TMB ox also serves as a near-infrared (NIR) photothermal agent to convert the colorimetric readout into heat under the irradiation of an 808 nm laser. A competitive-type immunoreaction is carried out between AFB 1 and glucose oxidase (GOx)-labeled AFB 1 -bovine serum albumin (AFB 1 -BSA-GOx) conjugates. With the formation of immune complexes, the entrained GOx catalyzes the hydrolysis of glucose to generate H 2 O 2, which further involves the Pt-CN catalyzed production of TMB ox to increase colorimetric/photothermal readouts. Depending on the degree of TMB oxidation, the dual-mode immunoassay provides a linear range of 1.0 pg/mL to 10 ng/mL, with a limit of detection (LOD) of 0.22 pg/mL for the colorimetric assay and 0.76 pg/mL for the photothermal assay. Moreover, the developed method is successfully used to detect AFB 1 in peanuts with acceptable accuracy compared with commercially enzyme-linked immunosorbent assay (ELISA) kits. Significantly, the photothermal readout in this method is recorded on a mobile phone device without any expensive instruments, providing an affordable and convenient tool for food safety testing. [ABSTRACT FROM AUTHOR]

NiCoBP-doped multi-walled carbon nanotube (NiCoBP–MWCNT) was first synthesized by using induced electroless-plating method and functionalized with the biomolecules for highly efficient electrochemical immunoassay of prostate-specific antigen (PSA, used as a model analyte). We discovered that the as-synthesized NiCoBP–MWCNT had the ability to catalyze the glucose oxidization with a stable and well-defined redox peak. The catalytic current increased with the increment of the immobilized NiCoBP–MWCNT on the electrode. Transmission electron microscope (TEM) and energy dispersive X-ray spectrometry (EDX) were employed to characterize the as-prepared NiCoBP–MWCNT. Using the NiCoBP–MWCNT-conjugated anti-PSA antibody as the signal-transduction tag, a new enzyme-free electrochemical immunoassay protocol could be designed for the detection of target PSA on the capture antibody-functionalized immunosensing interface. Experimental results revealed that the designed immunoassay system could exhibit good electrochemical responses toward target PSA, and allowed the detection of PSA at a concentration as low as 0.035 ng mL −1 . More importantly, the NiCoBP-MWCNT-based oxidase mimetic system could be further extended for the monitoring of other low-abundance proteins or disease-related biomarkers by tuning the target antibody. [ABSTRACT FROM AUTHOR]

Abstract: A novel class of molecular tags, cadmium ion-doped magnetic poly(styrene-acrylic acid) nanospheres (Cd-MPSA), was first synthesized and functionalized with polyclonal rabbit anti-human luteinizing hormone antibodies (PAb2) for highly efficient electrochemical immunoassay of luteinizing hormone (LH). Transmission electron microscope (TEM) and Fourier transform infrared spectroscope (FTIR) were employed to characterize the prepared Cd-MPSA. By using Cd-MPSA-labeled PAb2 as molecular tags, a novel sandwich-type immunoassay protocol was built for determination of LH on monoclonal mouse anti-human luteinizing hormone antibody (MAb1)-functionalized gold electrode. The assay was carried out in pH 5.3 HAc-NaAc buffer solution by square wave voltammetry (SWV). The signal was obtained by the reduction of the doped cadmium ions in the Cd-MPSA. Under optimal conditions, the currents increased with the increasing LH level in the sample, and exhibited a linear range from 0.25 to 240mIUmL−1 with a detection limit of 0.08mIUmL−1 LH at 3s B. The precision, reproducibility, and specificity were acceptable. No obvious difference was encountered in the analysis of spiking LH samples into newborn calf serum with the referenced values. [Copyright &y& Elsevier]

Abstract: A new electrochemical immunosensor for sensitive determination of thyroid-stimulating hormone (TSH) was designed by using redox-active nanogold-functionalized magnetic beads (GoldMag) as signal tags on the nanogold–graphene interface. To construct such GoldMag nanostructures, polyethyleneimine-functionalized magnetic beads (PEI-MBs) were initially prepared by using a wet chemical method, and the electroactive thionine molecules and gold nanoparticles were then alternately immobilized on the surface of PEI-MBs by using an opposite-charged adsorption technique and an in situ synthesis method, respectively. The synthesized GoldMag nanostructures were utilized as signal tags for the label of horseradish peroxidase-anti-TSH conjugates (HRP-anti-TSH). With a sandwich-type immunoassay format, the conjugated signal tags on the transducer were increased with the increasing TSH concentration in the sample, thus enhancing the signal of the electrochemical immunosensor due to the labeled HRP toward the catalytic reduction of H2O2. Under optimal conditions, the current was proportional to the logarithm of TSH concentration ranging from 0.01 to 20μIUmL−1 in pH 6.0 HAc–NaAc containing 6mM H2O2. The detection limit (LOD) was 0.005μIUmL−1 TSH at 3s B. The immunosensor displayed an acceptable reproducibility, stability and selectivity. In addition, the methodology was evaluated with human serum specimens, receiving good correlation with results from commercially available electrochemiluminescent analyzer. [Copyright &y& Elsevier]

Abstract: Methods based on sandwich-type immunosensors and immunoassays have been developed for detection of multivalent antigens/analytes with more than one eptiope due to the use of two matched antibodies. High-affinity antibodies and appropriate labels are usually employed for the amplification of detectable signal. Recent research has looked to develop innovative and powerful novel nanoparticle labels, controlling and tailoring their properties in a very predictable manner to meet the requirements of specific applications. This articles reviews recent advances, exploiting nanoparticle labels, in the sandwich-type immunosensors and immunoassays. Routine approaches involve noble metal nanoparticles, carbon nanomaterials, semiconductor nanoparticles, metal oxide nanostructures, and hybrid nanostructures. The enormous signal enhancement associated with the use of nanoparticle labels and with the formation of nanoparticle-antibody-antigen assemblies provides the basis for sensitive detection of disease-related proteins or biomolecules. Techniques commonly rely on the use of biofunctionalized nanoparticles, inorganic-biological hybrid nanoparticles, and signal tag-doped nanoparticles. Rather than being exhaustive, this review focuses on selected examples to illustrate novel concepts and promising applications. Approaches described include the biofunctionalized nanoparticles, inorganic-biological hybrid nanoparticles, and signal tage-doped nanoparticles. Further, promising application in electrochemical, mass-sensitive, optical and multianalyte detection are discussed in detail. [Copyright &y& Elsevier]

Abstract: Methods based on nanomaterial labels have been developed for electrochemical immunosensors and immunoassays, but most involved low sensitivity. Herein a novel class of molecular tags, nanogold–polyaniline–nanogold microspheres (GPGs), was first synthesized and functionalized with horseradish peroxidase-conjugated thyroid-stimulating hormone antibody (HRP-Ab2) for sensitive electrochemical immunoassay of thyroid-stimulating hormone (TSH). X-ray diffraction, confocal Raman spectroscopy, scanning electron microscope and transmission electron microscope were employed to characterize the prepared GPGs. Based on a sandwich-type immunoassay format, the assay was performed in pH 5.0 acetate buffer containing 6.0mmolL−1 H2O2 by using GPG-labeled HRP-Ab2 as molecular tags. Compared with pure polyaniline nanospheres and gold nanoparticles alone, the GPG hybrid nanostructures increased the surface area of the nanomaterials, and enhanced the immobilized amount of HRP-Ab2. Several labeling protocols comprising HRP-Ab2, nanogold particle-labeled HRP-Ab2, and polyaniline nanospheres-labeled HRP-Ab2, were also investigated for determination of TSH and improved analytical features were obtained by using the GPG-labeled HRP-Ab2. With the GPG labeling method, the effects of incubation time and pH of acetate buffer on the current responses of the immunosensors were also studied. The strong attachment of HRP-Ab2 to the GPGs resulted in a good repeatability and intermediate precision down to 7%. The dynamic concentration range spanned from 0.01 to 20μIUmL−1 with a detection limit (LOD) of 0.005μIUmL−1 TSH at the 3s B criterion. Significantly, no significant differences at the 0.05 significance level were encountered in the analysis of 15 spiking serum samples between the developed electrochemical immunoassay and the commercially available enzyme-linked immunosorbent assay (ELISA) method for determination of TSH. [Copyright &y& Elsevier]

Abstract: A novel class of redox-active molecular tags, poly(o-phenylenediamine)-carried nanogold particles (GPPDs), was first synthesized and functionalized with horseradish peroxidase-anti-prolactin conjugates (HRP-anti-PRL). Thereafter, a specific sandwich-type electrochemical immunoassay was designed for determination of prolactin (PRL) by using GPPD-labeled HRP-anti-PRL conjugates as molecular tags on anti-PRL antibody-modified glassy carbon electrode. Compared with pure gold nanoparticles and poly(o-phenylenediamine) microspheres, the as-prepared GPPDs increased the surface coverage of the nanostructures, and enhanced the immobilization amount of biomolecules. Several labeling protocols compromising GPPD-labeled HRP-anti-PRL, nanogold particles-labeled HRP-anti-PRL and poly(o-phenylenediamine) microspheres-labeled HRP-anti-PRL, were investigated for detection of PRL, and improved analytical features were obtained with the GPPD-based strategy. With the GPPD labeling method, dependence of the electrochemical signals on the incubation time and pH of the assay solution were also studied. The strong attachment of HRP-anti-PRL to the GPPDs resulted in a good repeatability and intermediate reproducibility down to 9.8%. The dynamic concentration range spanned from 0.5 to 180ngmL−1 PRL with a detection limit of 0.1ngmL−1 at the 3S blank level. No significant differences at the 95% confidence level were encountered in the analysis of 10 spiked blank cattle serum samples between the developed immunoassay and enzyme-linked immunosorbent assay method for determination of PRL. [Copyright &y& Elsevier]