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Chicken interleukin-4 (ChIL-4), which is released by activated type 2 helper (Th2) cells following their stimulation in vitro, is an important indicator for the study of cell-mediated immunity in chickens after infection or vaccination. In this work, the first ChIL-4 chemiluminescent (CL) immunosensor was developed via the immobilization of monoclonal ChIL-4 antibodies on a nitrogen-doped graphene (NG)-chitosan nanocomposite matrix. NG nanosheets were used for the first time in the CL immunoassay to provide a biocompatible microenvironment for the immobilized capture antibody. The ChIL-4 immunosensor was characterized systematically. The proposed immunosensor displayed a wide linear range from 0.05 to 70 ng mL −1 and a low detection limit of 0.02 ng mL −1 at a signal-to-noise ratio of 3. Compared to traditional assay methods, this system was more flexible, simple, rapid, and sensitive. Moreover, this CL immunoassay system had an excellent detection and fabrication reproducibility, a high specificity, an acceptable accuracy, and a high stability. This work enables the specific detection of ChIL-4 and the further study of its role in the immune responses of poultry. [ABSTRACT FROM AUTHOR]

Abstract: The quantitative detection of chicken interferon-γ (ChIFN-γ) released by T cells after in vitro stimulation is a good evaluation of cell-mediated immunity in chickens after infection or vaccination. This work reports a new flow-through the chemiluminescent (CL) immunoassay method for rapid and specific determination of ChIFN-γ at a level of sub-picogram per milliliter. The biocompatible graphene oxide nanosheet is introduced into CL immunoassay for highly efficient immobilization of capture antibody. The detection limit of the proposed method at a signal to noise ratio of 3 is 0.36pgml−1, which is 138-fold lower than the current lowest value of 50pgml−1 for ChIFN-γ. Coupling with a flow-through system, the whole immunoassay process can be completed within 25min. The resulting ChIFN-γ immunosensor shows excellent detection and fabrication reproducibility, good specificity and stability. The assay results of nature ChIFN-γ samples with the proposed method are in an acceptable agreement with the reference values. Compared to the present assay methods, this method is more flexible, simple, rapid and sensitive. The aim of this work is to demonstrate that the ultrasensitive, specific and rapid CL immunoassay format can become a very potential application for quantifying ChIFN-γ and further studying its role in immune response in poultry. [Copyright &y& Elsevier]

Highlights: [•] An ultrasensitive chemiluminescent (CL) immunoassay strategy was developed. [•] This assay was preformed based on a streptavidin-functionalized MWCNTs platform. [•] The sensitivity displays 7.9-fold increase compared to that without MWCNTs. [•] This method provides a promising approach for ultrasensitive CL immunoassay. [Copyright &y& Elsevier]

Abstract: A streptavidin functionalized capillary immune microreactor was designed for highly efficient flow-through chemiluminescent (CL) immunoassay. The functionalized capillary could be used as both a support for highly efficient immobilization of antibody and a flow cell for flow-through immunoassay. The functionalized inner wall and the capture process were characterized using scanning electron microscopy. Compared to conventional packed tube or thin-layer cell immunoreactor, the proposed microreactor showed remarkable properties such as lower cost, simpler fabrication, better practicality and wider dynamic range for fast CL immunoassay with good reproducibility and stability. Using α-fetoprotein as model analyte, the highly efficient CL flow-through immunoassay system showed a linear range of 3 orders of magnitude from 0.5 to 200ngmL−1 and a low detection limit of 0.1ngmL−1. The capillary immune microreactor could make up the shortcoming of conventional CL immunoreactors and provided a promising alternative for highly efficient flow-injection immunoassay. [Copyright &y& Elsevier]

Abstract: A novel immunoaffinity column used as an immunosensor for flow-injection chemiluminescent (CL) immunoassay was prepared by immobilizing antibody on carboxylic resin beads. The immunosensor could fast recognize and trap the immunocomplex of horseradish peroxidase (HRP)-labeled antibody and antigen, which was firstly formed with a micro-bubble accelerated pre-incubation process, to produce a sandwich immunocomplex. The HRP introduced in the immunoaffinity column could catalyze the CL reaction to produce enzyme-enhanced emission. With α-fetoprotein (AFP) as a mode, a flow-injection CL immunoassay was proposed. The whole assay for one sample, including the pre-incubation and the regeneration of immunoaffinity column, could be performed within 16min. The linear range was 1.0–80ng/ml with a correlation coefficient of 0.998 and a detection limit of 0.1ng/ml at a signal/noise ratio of 3. The intra- and inter-assay coefficients of variation at 20ng/ml AFP were 1.2% and 8.5%, respectively. The storage stability of the immunoaffinity column and the accuracy for sample detection were acceptable. This flexible, sensitive, low-cost, and rapid method is valuable for clinical immunoassay. [Copyright &y& Elsevier]

Abstract Quantitative detection of multiple chicken cytokines is a good evaluation of cell-mediated immunity in chickens after disease infection or vaccination. However, current assay methods for chicken cytokines cannot meet the needs of clinical diagnosis due to unsatisfactory sensitivity and low assay throughput. Herein, a sensitive chemiluminescence (CL) imaging immunosensor array has been developed for high-throughput detection of multiple chicken cytokines. The chicken cytokines immunosensor array was prepared by assembling different cytokine capture antibodies onto a disposable silanized glass chip, where horseradish peroxidase and antibody-conjugated gold nanoparticles were used as multienzymatic amplification probe for CL imaging signal amplification. By using a sandwich assay mode, the amplified CL signals from each sensing array cell were collected for quantitation. Using chicken interleukin-4 and chicken interferon-γ as model cytokines, this novel multiplexed and amplified method demonstrated simultaneous measurement of the two chicken cytokines in the linear ranges of 0.008–0.12 ng/mL and 0.005–0.20 ng/mL, respectively, which yields limits of detection down to 2 pg/mL and 3 pg/mL. The CL imaging array method reported here also demonstrated high specificity, good repeatability, and high stability and accuracy, providing a novel multiplex immunoassay strategy for highly sensitive and high-throughput detection of chicken cytokines and further disease diagnosis in poultry. Graphical abstract A novel multiplex immunosensor array for sensitive and high-throughput detection of multiple chicken cytokines is for the first time developed based on highly-sensitive chemiluminescence imaging coupled with a multienzymatic signal amplification probe. Image 1 Highlights • Multiplex immunosensor array for chicken cytokines was proposed for the first time. • Multiple chicken cytokines were simultaneously detected by sensitive CL imaging system. • The multiplex cytokines immunoassay has much better performance compared with current methods. • This work provides a novel strategy for sensitive and high-throughput multiplex immunoassay. [ABSTRACT FROM AUTHOR]

Dual-functional cupric oxide nanorods (CuONRs) as peroxidase mimics are proposed for the development of a flow-through, label-free chemiluminescent (CL) immunosensor. Forming the basis of this cost-efficient, label-free immunoassay, CuONRs, synthesized using a simple hydrothermal method, were deposited onto epoxy-activated standard glass slides, followed by immobilization of biotinylated capture antibodies through a streptavidin bridge. The CuONRs possess excellent catalytic activity, along with high stability as a solid support. Antigens could then be introduced to the sensing system, forming large immunocomplexes that prevent CL substrate access to the surface, thereby reducing the CL signal in a concentration dependent fashion. Using carcinoembryonic antigen (CEA) as a model analyte, the proposed label-free immunosensor was able to rapidly determine CEA with a wide linear range of 0.1–60 ng mL −1 and a low detection limit of 0.05 ng mL −1 . This nanozyme-based immunosensor is simple, sensitive, cost-efficient, and has the potential to be a very promising platform for fast and efficient biosensing applications. [ABSTRACT FROM AUTHOR]

A novel photoelectrochemical immunosensor was constructed to monitor carcinoembryonic antigen (CEA) based on hybridization chain reaction (HCR)-mediated in situ generation of copper nanoparticles (Cu NPs) and subsequent Cu2+-dependent quenching reaction, in which titanium dioxide nanoparticles-sensitized double-shell zinc cadmium sulfide hollow nanospheres (TiO 2 /DS-ZnCdS)-modified ITO electrode and anti-CEA antibody-modified 96-well plate served as biological recognition and signal detection platforms, respectively. The synergistic effect of TiO 2 NPs and DS-ZnCdS hollow nanospheres contributed to the improvement of photoelectric conversion efficiency, and HCR-mediated signal cascade benefited the enhancement of detection sensitivity. In the presence of CEA, biotin-labelled anti-CEA antibodies were immobilized onto anti-CEA antibody-modified 96-well plate, and triggered HCR process to form long double stranded DNA, which could adsorb a large number of Cu2+ ions and then in situ form Cu NPs on double stranded DNA template by a facile reduction reaction. After acid treatment, the dissolved Cu2+ ions could significantly reduce the photocurrent response due to the generation of Cu x S. Under optimal conditions, the immunosensor exhibited a desirable liner range of 1 pg mL−1 – 50 ng mL−1 and a low detection limit of 0.1 pg mL−1, as well as excellent selectivity and stability. Meanwhile, the recoveries of human serum sample analysis ranged from 96.8% to 103.6%, and the relative standard deviation was less than 7.40%, showing a good feasibility in early clinical diagnosis. • A novel photoelectrochemical immunosensor based on TiO 2 -sensitized DS-ZnCdS HNs was constructed for CEA detection. • HCR-mediated cascade process and Cu2+-based quenching reaction were integrated. • The photocurrent response was enhanced by TiO 2 /DS-ZnCdS heterostructure. • The excellent performances of designed immunosensor benefited to its potential application in early clinical diagnosis. [ABSTRACT FROM AUTHOR]