Electrochemical Signal Tracing by Glucose Oxidase and Ferrocene Dually Functionalized Gold Nanoprobe for Ultrasensitive Immunoassay

A glucose oxidase (GOD) and ferrocene (Fc) dually functionalized gold nanoprobe was simply prepared for electrochemical immunoassay. By combination with sandwich immunoreaction at a carbon nanotube (CNT)‐based immunosensor and signal tracing of the nanoprobe through the Fc‐mediated GOD‐catalytic reaction, a new electrochemical immunoassay method was successfully developed. Both the multi‐enzyme signal amplification of the nanoprobe and the electron transfer promotion of the CNTs modified on the immunosensor greatly enhanced the signal response. Thus this method showed excellent analytical performance including ultrahigh sensitivity, wide linear range as well as good specificity, reproducibility, stability and reliability for human IgG measurement.     

双重信号放大的竞争型免疫传感器的制备及其应用于瘦肉精检测的研究

制备了葡萄糖氧化酶(GOD)-克伦特罗(Clenbuterol, CB)功能纳米复合物, 并采用共价键合和温育组装等方法构建了双重信号放大的竞争型免疫传感器. 研究了GOD 催化氧化葡萄糖和普鲁士蓝(PB)催化还原H2O2 双重信号放大的反应机理和传感器检测CB 的作用机制. 用扫描电子显微镜(SEM)等方法表征了纳米复合材料的形貌和复合物中GOD的活性, 复合物中的GOD 保持了良好的电催化性能和酶动力学响应, 并且符合米氏动力学方程. 最佳实验条件下, 该免疫传感器对盐酸克伦特罗的检测线性范围为0.01～100 ng/mL, 检测限达4.50 pg/mL. 实验结果表明, 该传感器对瘦肉精克伦特罗的检测具有灵敏度高, 特异性强, 重现性好, 线性范围宽和检测限低等优点. 将该方法用于猪肝样品的分析, 加标样品回收率在97.5%～102%之间. 该研究为瘦肉精及β-受体兴奋剂的分析提供了一种新方法.     

Dual‐Amplification of Antigen–Antibody Interactions via Backfilling Gold Nanoparticles on (3‐Mercaptopropyl) Trimethoxysilane Sol‐Gel Functionalized Interface

A new dual‐amplification strategy of electrochemical signaling from antigen–antibody interactions was proposed via backfilling gold nanoparticles on (3‐mercaptopropyl) trimethoxysilane sol‐gel (MPTS) functionalized interface. The MPTS was employed not only as a building block for the electrode surface modification but also as a matrix for ligand functionalization with first amplification. The second signal amplification strategy introduced in this study was based on the backfilling immobilization of nanogold particles to the immunosensor surface. Several coupling techniques, such as with nanogold but not MPTS or with MPTS but not nanogold, were investigated for the determination of carcinoembryonic antigen (CEA) as a model, and a very good result was obtained with nanogold and MPTS coupling immunosensor. With the noncompetitive format, the formation of the antigen–antibody complex by a simple one‐step immunoreaction between the immobilized anti‐CEA and CEA in sample solution introduced membrane potential change before and after the antigen–antibody interaction. Under optimal conditions, the proposed immunosensor exhibited a good electrochemical behavior to CEA in a dynamic concentration range of 4.4 to 85.7 ng/mL with a detection limit of 1.2 ng/mL (at 3 δ). Moreover, the precision, reproducibility and stability of the as‐prepared immunosensor were acceptable. Importantly, the proposed methodology would be valuable for diagnosis and monitoring of carcinoma and its metastasis.     

Polymerization-assisted signal amplification for electrochemical detection of biomarkers

We present a novel immunosensor by using polymerization-assisted signal amplification strategy coupled with electrochemical detection. A sandwich immunoassay process was used to immobilize a polymerization reaction center, the initiator-conjugated polyclonal prostate specific antigen (PSA) or polyclonal carcinoembryonic antigen (CEA) antibodies on the surface of the electrode. Activator generated electron transfer for atom transfer radical polymerization (AGET ATRP) subsequently triggered the local accumulation of glycidyl methacrylate (GMA) monomers. Growth of long chain polymers provided excess epoxy groups for electrochemical tags aminoferrocene (FcNH(2)) coupling, which in turn significantly increased the loading of the signal molecules and enhanced the electrochemical readouts. The detection limit was ～0.14 pg mL(-1) for PSA and ～0.10 pg mL(-1) for CEA in PBS buffers. The proposed immunosensor was highly sensitive, selective and has a good match to the clinical electrochemiluminescent method. This suggested that the polymerization-assisted immunosensing strategy could be used as an effective method to significantly enhance signal output of the sandwich immunoassays and acted as a promising platform for the clinical screening of cancer biomarkers.     

Ultrasensitive electrochemical immunosensor of carcinoembryonic antigen based on gold-label silver-stain signal amplification

A novel gold-label silver-stain electrochemical immunosensor based on polythionine-gold nanoparticles (PTh-Au NPs) modified glassy carbon electrode (GCE) as a platform and secondary antibody labeled Au NPs (Ab₂-Au NPs) as immumoprobe for carcinoembryonic antigen (CEA) detection. The sandwich-type biosensor adopted anodic stripping voltammetry to detect silver stripping signal when the Ab₂-Au NPs of the formed immunocomplexes were stained with silver.     

Amplified inhibition of the electrochemical signal of ferrocene by enzyme-functionalized graphene oxide nanoprobe for ultrasensitive immunoassay

A nanoprobe-induced signal inhibition mechanism was designed for ultrasensitive electrochemical immunoassay at a chitosan-ferrocene (CS-Fc) based immunosensor. The nanoprobe was prepared by covalently loading signal antibody and high-content horseradish peroxidase (HRP) on the graphene oxide (GO) nanocarrier. The immunosensor was prepared through the stepwise assembly of gold nanoparticles (Au NPs) and capture antibody at a CS-Fc modified electrode. After sandwich immunoreaction, the GO-HRP nanoprobes were quantitatively captured onto the immunosensor surface and thus induced the production of a layer of insoluble film through the enzymatically catalytic reaction of the HRP labels. Both the dielectric immunocomplex formed on the immunosensor surface and the enzymatic precipitate with low electroconductivity led to the electrochemical signal decease of the Fc indicator, which was greatly amplified by the multi-enzyme signal amplification of the nanoprobe. Based on this amplified signal inhibition mechanism, a new ultrasensitive electrochemical immunoassay method was developed. Using carcinoembryonic antigen as a model analyte, this method showed a wide linear range over 5 orders of magnitude with a detection limit down to 0.54 pg/mL. Besides, the immunosensor showed good specificity, acceptable reproducibility and stability as well as satisfactory reliability for the serum sample analysis.     

Sensitive electrochemical immunosensor array for the simultaneous detection of multiple tumor markers

A novel electrochemical immunosensor array for the simultaneous detection of multiple tumor markers was developed by incorporating electrochemically addressing immobilization and one signal antibody strategy. As a proof-of-principle, an eight-electrode array including six carbon screen-printed working electrodes was used as a base array for the analysis of two important tumor markers, carcinoembryonic antigen (CEA) and α-fetoprotein (AFP) and a horseradish peroxidase-labeled antibody was employed as a signal antibody. The immunosensor in the array was fabricated in sequence by covalently coupling the capture antibody onto the surface of the desired working electrode, which was firstly electrochemically addressably grafted with an aminophenyl group by reduction of in situ generated aminophenyl diazonium cation generated from p-phenylenediamine, using glutaraldehyde as cross-linker. This allowed the selective immobilization of the capture antibody at the desired position on a single array via an electrochemical operation. The immunoassay in sandwich mode was performed by specifically binding the targets, second antibodies and one signal antibody to the immunosensor array. The result showed that the steady current density was directly proportional to the concentration of target CEA/AFP in the range from 0.10 to 50 ng mL(-1) with a detection limit of 0.03 ng mL(-1) for CEA and 0.05 ng mL(-1) for AFP (S/N = 3), respectively. This work demonstrates that the employment of an electrochemically addressing method for the fabrication of an immunosensor array and one signal antibody is a promising approach for the determination of multiple tumor markers in clinical samples.     

Disposable electrochemical immunosensor by using carbon sphere/gold nanoparticle composites as labels for signal amplification

This work designed a simple, sensitive, and low-cost immunosensor for the detection of protein marker by using a carbon sphere/gold nanoparticle (CNS/AuNP) composite as an electrochemical label. The nanoscale carbon spheres, prepared with a hydrothermal method by using glucose as raw material, were used to load AuNPs for labeling antibody by electrostatic interaction, which provided a feasible pathway for electron transfer due to the remarkable conductivity. The disposable immunosensor was constructed by coating a polyethylene glycol (PEG) film on a screen-printed carbon-working electrode and then immobilizing capture antibody on the film. With a sandwich-type immunoassay format, the analyte and then the CNS/AuNP-labeled antibody were successively bound to the immunosensor. The bound AuNPs were finally electro-oxidized in 0.1 M HCl to produce AuCl(4)(-) for differential pulse voltammetric (DPV) detection. The high-loading capability of AuNPs on CNS for the sandwich-type immunorecognition led to obvious signal amplification. By using human immunoglobulin?G (IgG) as model target, the DPV signal of AuNPs after electro-oxidized at optimal potential of +1.40?V for 40?s showed a wide linear dependence on the logarithm of target concentration ranging from 10?pg mL(-1) to 10?ng mL(-1). The detection limit was around 9?pg mL(-1). The immunosensor showed excellent analytical performance with cost effectivity, good fabrication reproducibility, and acceptable precision and accuracy, providing significant potential application in clinical analysis.     

The Signal‐Enhanced Label‐Free Immunosensor Based on Assembly of Prussian Blue‐SiO2 Nanocomposite for Amperometric Measurement of Neuron‐Specific Enolase

A signal‐enhanced label‐free electrochemical immunosensor was constructed by the employment of Prussian blue doped silica dioxide (PB‐SiO₂) nanocomposite. At first, PB‐SiO₂ nanocomposite which was produced by using a microemulsion method was used to obtain a nanostructural monolayer on a glassy carbon electrode (GCE) surface. Next amino‐functionalized interface were prepared by self‐assembling 3‐aminopropyltriethoxy silane (APTES) on the PB‐SiO₂ nanoparticle surface. Then chitosan stabled gold nanoparticle (CS‐nanoAu) was subsequently attached, while the entire surface was finally loaded with neuron‐specific enolase antibody (anti‐NSE) via the adsorption of gold nanoparticle. The sensitivity of the proposed immunosensor has greatly improved as the PB‐SiO₂ nanostructural sensing film provides plenty of active sites which might catalyze the reduction of H₂O₂. The immunosensor exhibited good linear behavior in the concentration range from 0.25–5.0 and 5.0–75 ng/mL for the quantitative analysis of neuron‐specific enolase (NSE), a putative serum marker of small‐cell lung carcinoma (SCLC), with a limit of detection of 0.08 ng/mL. The resulting NSE immunosensor showed high sensitivity and long‐term lifetime which can be attributed to the extremely high catalytic activity and biocompatibility of CS‐nanoAu/APTES/PB‐SiO₂ nanostructural multilayers.     

基于花状铂纳米颗粒构建的电致化学发光免疫传感器用于检测载脂蛋白A1

采用一锅合成法制备了新型的具有大比表面积的花状铂纳米颗粒（PtNFs）,并构建了一个高灵敏电致化学发光（ECL）免疫传感器用于检测载脂蛋白A1（Apo-A1）. 该PtNFs用于吸附二抗（anti-Apo-A1）,并用葡糖糖氧化酶（GOD）封闭其表面的非特异性位点,最终制备了PtNFs@anti-Apo-A1@GOD信号探针. 当Apo-A1存在时,通过夹心免疫反应将制备的信号探针捕获于电极表面,并将所制得的电极置于含有葡萄糖的过硫酸根底液中检测. GOD催化葡萄糖产生H₂O₂,H₂O₂在PtNFs的催化下分解并在电极表面原位产生O₂,所产生的O₂能够催化过硫酸根-氧气体系的电致化学发光反应,放大发光信号,提高检测灵敏度. 该传感器在0.1ng•mL^-1 ~ 100 ng•mL^-1范围内对Apo-A1有良好的线性响应,检测下限达到0.03ng•mL^-1,有望应用于临床分析诊断.     

A reusable electrochemical immunosensor for carcinoembryonic antigen via molecular recognition of glycoprotein antibody by phenylboronic acid self-assembly layer on gold

A reusable amperometric immunosensor based on the reversible boronic acid-sugar interaction is proposed. The immunosensor was prepared by self-assembling a thiol-mixed monolayer comprised of conjugates of 3-aminophenylboronic acid with 11-mercaptoundecanoic acid (APBA-MUA) and 11-mercapto-1-undecanol (MU) on gold. The resulting boronic acid coating layer can specifically bind with the glycoprotein antibody, enzyme conjugated carcinoembryonic antibody (HRP-anti-CEA). Voltammetric and electrochemical impedance spectroscopic (EIS) studies and surface plasmon resonance (SPR) measurements show that the binding of HRP-anti-CEA to the APBA interface is reversible and the HRP-anti-CEA can be removed with an acidic buffer or a solution containing sorbitol. The bound enzyme-conjugated antibody can retain its enzyme catalytic activity to the reduction of hydrogen peroxide (H(2)O(2)) and its immunoactivity while binding with CEA to form an immunocomplex. After the formation of the immunocomplex, the access of the active center of HRP to thionine was partially inhibited. This leads to a linear decrease in the electrocatalytic response of HRP-anti-CEA-modified electrode over a CEA concentration range of 2.5 to 40.0 ng mL(-1). After monitoring the immunoreaction signals, the immunocomplex can be easily removed from the APBA interface with a regeneration solution. This regenerated APBA interface can rebound with HRP-anti-CEA and be recognized by the antigen, through which a reusable immunosensor with an RSD of 7.1% for four cycles can be obtained. Under optimal conditions, the detection limit for the CEA immunoassay is 1.1 ng mL(-1), at three times background noise. Serum CEA determination results, obtained with the proposed method, shows that the immunosensor has an acceptable accuracy.     

An ultrasensitive luminol cathodic electrochemiluminescence immunosensor based on glucose oxidase and nanocomposites: Graphene–carbon nanotubes and gold-platinum alloy

In the present study, a novel and ultrasensitive electrochemiluminescence (ECL) immunosensor based on luminol cathodic ECL was fabricated by using Au nanoparticles and Pt nanoparticles (nano-AuPt) electrodeposited on graphene–carbon nanotubes nanocomposite as platform for the detection of carcinoembryonic antigen (CEA). For this introduced immunosensor, graphene (GR) and single wall carbon nanotubes (CNTs) dispersed in chitosan (Chi-GR-CNTs) were firstly decorated on the bare gold electrode (GE) surface. Then nano-AuPt were electrodeposited (DpAu-Pt) on the Chi-GR-CNTs modified electrode. Subsequently, glucose oxidase (GOD) was employed to block the non-specific sites of electrode surface. When glucose was present in the working buffer solution, GOD immediately catalyzed the oxidation of glucose to in situ generate hydrogen peroxide (H₂O₂), which could subsequently promote the oxidation of luminol with an amplified cathodic ECL signal. The proposed immunosensor was performed at low potential (−0.1 to 0.4 V) and low concentration of luminol. The CEA was determined in the range of 0.1 pg mL⁻¹ to 40 ng mL⁻¹ with a limit of detection down to 0.03 pg mL⁻¹ (S N⁻¹ = 3). Moreover, with excellent sensitivity, selectivity, stability and simplicity, the as-proposed luminol-based ECL immunosensor provided great potential in clinical applications.     

磁性纳米金共价固定癌胚抗原单克隆抗体的电流型免疫传感器

合成了Fe3O4/Au磁性复合纳米粒子, 在粒子表面通过自组装硫脲分子使表面氨基化, 再用戊二醛共价交联固定癌胚抗原抗体(anti-CEA). 在外加磁场的作用下, 将anti-CEA复合磁性粒子吸附在固体石蜡碳糊电极表面, 制成了新型电流型免疫传感器. 免疫电极在含有癌胚抗原CEA和辣根过氧化物酶标记的癌胚抗原(HRP-CEA)的混合溶液中温育, CEA和HRP-CEA与固定在电极表面的anti-CEA发生竞争反应, 导致HRP对H2O2的催化降解作用的改变, 从而可间接测定CEA. 由于标记的HRP可催化降解H2O2, 导致媒介体间苯二酚浓度改变, 使测定的灵敏度大大提高. 响应电流与CEA质量浓度的对数在2~160 ng/mL的范围内呈线性关系, 检出限为0.57 ng/mL(3σ法). 该免疫传感器具有制作简单、价廉及表面易于更新等特点.     

Dual amplification strategy for the fabrication of highly sensitive interleukin-6 amperometric immunosensor based on poly-dopamine

An electrochemical immunosensor was studied for sensitive detection of Interleukin-6 (IL-6) based on a dual amplification mechanism resulting from Au nanoparticles (AuNP)-Poly-dopamine (PDOP) as the sensor platform and multienzyme-antibody functionalized AuNP-PDOP@carbon nanotubes (CNT). The stable and robust film, PDOP, was used to immobilize biomolecules not only for the construction of the sensor platform, but also for the signal labeling. Sensitivity was greatly amplified by using the special platform of AuNP-PDOP and synthesizing horseradish peroxidase (HRP)-antibody (Ab(2)) functionalized AuNP-PDOP@carbon nanotubes (CNT). A linear response range of IL-6 from 4.0 to 8.0 × 10(2) pg mL(-1) with a low detection limit of 1.0 pg mL(-1) was obtained by the amperometry determination. Measurements of IL-6 in human serum gave excellent correlations with standard ELISA assays. Moreover, the immunosensor exhibited high selectivity, good reproducibility, and stability.     

A novel lable-free electrochemical immunosensor for carcinoembryonic antigen based on gold nanoparticles-thionine-reduced graphene oxide nanocomposite film modified glassy carbon electrode

In this paper, gold nanoparticle-thionine-reduced graphene oxide (GNP-THi-GR) nanocomposites were prepared to design a label-free immunosensor for the sensitive detection of carcinoembryonic antigen (CEA). The nanocomposites with good biocompatibility, excellent redox electrochemical activity and large surface area were coated onto the glassy carbon electrode (GCE) surface and then CEA antibody (anti-CEA) was immobilized on the electrode to construct the immunosensor. The morphologies and electrochemistry of the formed nanocomposites were investigated by using scanning electron microscopy (SEM), ultraviolet-visible (UV-vis) spectrometry, electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). CV and differential pulse voltammetry (DPV) studies demonstrated that the formation of antibody-antigen complexes decreased the peak current of THi in the GNP-THi-GR nanocomposites. The decreased currents were proportional to the CEA concentration in the range of 10-500 pg/mL with a detection limit of 4 pg/mL. The proposed method was simple, fast and inexpensive for the determination of CEA at very low levels.     

A novel electrochemical immunosensor using β-cyclodextrins functionalized silver supported adamantine-modified glucose oxidase as labels for ultrasensitive detection of alpha-fetoprotein

In this work, a novel sandwich-type electrochemical immunosensor based on host-guest interaction was fabricated for the detection of alpha-fetoprotein (AFP). Due to the large specific surface area of multiwalled carbon nanotubes and the unique supramolecular recognition ability of β-cyclodextrins, ferrocenecarboxylic acid (Fc) was incorporated into this sensor platform by host-guest interaction to generate an electrochemical signal. And β-cyclodextrins functionalized silver supported adamantine-modified glucose oxidase (GOD-CD-Ag), was used as a label to improve the analytical performance of the immunosensor by the dual amplification strategy. The obtained GOD-CD-Ag conjugates could convert glucose into gluconic acid with the formation of hydrogen peroxide (H₂O₂). And then silver nanoparticles could in situ catalyze the reduction of the generated H₂O₂, dramatically improving the oxidation reaction of Fc. The developed immunosensor shows a wide linear calibration range from 0.001 to 5.0 ng/mL with a low detection limit (0.2 pg/mL) for the detection of AFP. The method, with ideal reproducibility and selectivity, has a wide application prospect in clinical research.     

Fabrication of Prussian Blue modified ultramicroelectrode for GOD imaging using scanning electrochemical microscopy

A Prussian Blue (PB) film modified disk ultramicroelectrode (UME) was fabricated by electrochemical deposition technique on a Pt-disk UME. The electrocatalytical reductions of hydrogen peroxide derived from glucose oxidase (GOD) on this modified UME were investigated. The enzymatic biochemical reactivity was imaged by scanning electrochemical microscopy (SECM) utilizing the PB film modified UME. It is evident that sensitivity and spatial resolution for hydrogen peroxide measurement were improved obviously. SECM images obtained clearly revealed the concentration profile of the reaction products around the enzymes. The PB film modified microelectrode is in the nature of simple preparation, high catalytic activity on hydrogen peroxide and substrate selectivity for SECM etc.     

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.     

Immobilized Fullerene C60‐Enzyme‐Based Electrochemical Glucose Sensor

A mixed‐valence cluster of cobalt(II) hexacyanoferrate and fullerene C60‐enzyme‐based electrochemical glucose sensor was developed. A water insoluble fullerene C60‐glucose oxidase (C60‐GOD) was prepared and applied as an immobilized enzyme on a glassy carbon electrode with cobalt(II) hexacyanoferrate for analysis of glucose. The glucose in 0.1 M KCl/phosphate buffer solution at pH = 6 was measured with an applied electrode potential at 0.0 mV (vs Ag/AgCl reference electrode). The C60‐GOD‐based electrochemical glucose sensor exhibited efficient electro‐catalytic activity toward the liberated hydrogen peroxide and allowed cathodic detection of glucose. The C60‐GOD electrochemical glucose sensor also showed quite good selectivity to glucose with no interference from easily oxidizable biospecies, e.g. uric acid, ascorbic acid, cysteine, tyrosine, acetaminophen and galactose. The current of H₂O₂ reduced by cobalt(II) hexacyanoferrate was found to be proportional to the concentration of glucose in aqueous solutions. The immobilized C60‐GOD enzyme‐based glucose sensor exhibited a good linear response up to 8 mM glucose with a sensitivity of 5.60 × 10² nA/mM and a quite short response time of 5 sec. The C60‐GOD‐based glucose sensor also showed a good sensitivity with a detection limit of 1.6 × 10^‐6 M and a high reproducibility with a relative standard deviation (RSD) of 4.26%. Effects of pH and temperature on the responses of the immobilized C60‐GOD/cobalt(II) hexacyanoferrate‐based electrochemical glucose sensor were also studied and discussed.     

Ultrasensitive electrochemiluminescence immunoassay for tumor marker detection using functionalized Ru-silica@nanoporous gold composite as labels

In this work, we reported a simple and sensitive sandwich-type electrochemiluminescence (ECL) immunosensor for carcinoembryonic antigen (CEA) on a gold nanoparticles (AuNPs) modified glassy carbon electrode (GCE). The Ru-silica (Ru(bpy)(3)(2+)-doped silica) capped nanoporous gold (NPG) (Ru-silica@NPG) composite was used as an excellent label with amplification techniques. The NPG was prepared with a simple dealloying strategy, by which silver was dissolved from silver/gold alloys in nitric acid. The primary antibody was immobilized on the AuNPs modified electrode through l-cysteine and glutaraldehyde, and then the antigen and the functionalized Ru-silica@NPG composite labeled secondary antibody were conjugated successively to form a sandwich-type immunocomplex through the specific interaction. The concentrations of CEA were obtained in the range from 1 pg mL(-1) to 10 ng mL(-1) with a detection limit of 0.8 pg mL(-1). The as-proposed ECL immunosensor has the advantages of high sensitivity, specificity and stability and could become a promising technique for tumor marker detection.