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High-sensitivity electrochemiluminescence probe with molybdenum carbides as nanocarriers for #-fetoprotein sensing Xiaoqing Zhu, Qingfeng Zhai, Wenling Gu, Jing Li, and Erkang Wang Anal. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.analchem.7b02701 • Publication Date (Web): 26 Oct 2017 Downloaded from http://pubs.acs.org on October 26, 2017

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Analytical Chemistry

High-sensitivity electrochemiluminescence probe with molybdenum carbides as nanocarriers for α-fetoprotein sensing Xiaoqing Zhu,[a,b] Qingfeng Zhai,[a,c] Wenling Gu,[a,c] Jing Li,*[a] and Erkang Wang*[a] [a] State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun, Jilin, 130022, P. R. China

[b] University of Science and Technology of China, Hefei, Anhui, 230026, P. R. China

[c] University of the Chinese Academy of Sciences, Beijing, 100039, P. R. China

*Corresponding author: Assoc. Prof. Jing Li and Prof. Erkang Wang, Tel: +86-431-85262003, Email: [email protected] and [email protected]

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ABSTRACT

Suitably designed electrochemiluminescence (ECL) carrying group acting as high-efficiency solid-state probe has attracted a lot of attention. Herein, molybdenum carbides with the two-dimensional ultrathin nanosheet structure on the surface and excellent conductivity were successfully employed as the nanocarriers for the capture of ECL reagent of luminol capped Au nanoparticles (luminol-AuNPs). Notably, the luminol-AuNPs in the hybrid (luminol-AuNPs@Mo2C) exhibited enhanced ECL performance (ca. 6-fold) as compared to individual luminol-AuNPs due to the facilitated electron transfer process. Ultimately, the as-prepared ECL label was used to construct a label-free ECL immunosensor for the detection of α-fetoprotein (AFP). The immunosensor shows high selectivity and high sensitivity to AFP detection with a wide linear range of 0.1 pg·mL-1 to 30 ng·mL-1 and an extremely low detection limit of 0.03 pg·mL-1 (S/N=3). Moreover, the fabricated ECL immunosensor exhibit satisfied performance in the practical application. This novel sensing strategy not only broadens the application of molybdenum carbides but also provides a new efficient approach to detect various biomolecules.


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INTRODUCTION Electrochemiluminescence (ECL) is a light emitting progress triggered by electron-transfer reaction to form excited states.1,2 ECL is the marriage of chemiluminescence and electrochemistry, which endows it with many inherent advantages such as versatility, high sensitivity, wide dynamic range, potential and spatial control, simplified optical setup and low background signals.3-8 Since the first detailed investigation reported in mid-1960s,

9,10

ECL has been attracted more and more attentions and

has been applied in various areas including immunoassays,11 DNA probe assays,12 cell imaging13. The luminol as an ECL luminophore has become more popular owing to its high efficiency, low cost, good chemical stability and low oxidation potential.14,15 However, most of the luminol-based ECL system was performed in the solution which would make the emission efficiency discount greatly.16 Therefore, suitably designed ECL carrying group for improving the ECL efficiency is very important, especially for its analytical and bioanalytical applications. One effective strategy was based on the functionalized nanomaterial as carriers due to the unique electronic, optical and catalytical features.17,18 For instance, Cui’s group demonstrated that the luminol could be acted as capping ligands of Au nanoparticles (luminol-AuNPs) to fabricate the solid-state ECL probe with high efficiency due to the large surface area and excellent catalytic effect of Au nanoparticles.19,20 However, the ECL signals of the luminol-AuNPs were weak and unstable after conjugated with antibody,21 which limited the further application of individual luminol-AuNPs for immunosensor. To further improve the ECL behavior, nanomaterials with excellent electron-transfer and catalytical capability were often introduced into the luminol-AuNPs system (eg. graphene, multiwalled carbon nanotubes and Au nanoparticles).21-23 Despite these advances, the synthesis of high-efficiency ECL probe is still a great challenge, mainly because of the difficulty of combining excellent electron transfer properties with enhanced ECL efficiency simultaneously. In recent years, transition metal carbides (TMCs) have attracted growing research interest due to their outstanding physical and chemical properties benefiting from the unique metal-C chemical bonding and Pt-like d-state density around the Fermi level, such as exceptional hardness, thermal stability, high electric conductivity and resistance against corrosion.24-28 Especially, nanostructured molybdenum carbides (Mo2C) are considered as the most promising TMCs and have been applied in many fields including Li-ion batteries,29 dye-sensitized solar cells,30 supercapacitors,31 water-gas shift reaction32 and ACS Paragon Plus Environment


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electrocatalytic reactions33,34 (oxygen reduction reaction and hydrogen evolution reaction). For example, Zhu’s group synthesized mesoporous Mo2C/N-doped carbon heteronanowires and used as anode material for Li-ion battery, which exhibited dramatically capacity and rate capability, as well as long-term cycle life.35 In addition, Mo2C is also explored as high efficient hydrogen evolution reaction catalyst and considered as the most promising alternative to platinum owing to its low cost, high chemical stability, excellent electrical conductivity and high similarity to Pt-based catalyst.

28,36

Although, Mo2C is the popular material in energy storage and conversion, few works have been reported it in the construction of biosensor.37,38 For example, the label-free electrochemical determination of AFP with high sensitivity was constructed in our previous work with thionin capped Mo2C nanotube, where the Mo2C nanotube was used as carriers to load thionin electrochemical probes.37 In addition, based on the different adsorption ability of Mo2C to ss-DNA and ds-DNA, the fluorescence assay was also developed for the bisphenol A detection.38 However, the use of Mo2C as the carriers for loading ECL luminophore has so far not been reported. Given the tremendous electronic conductivity and efficient catalytic efficiency,39 the introduction of Mo2C will be indeed a promising route for enhanced efficiency. In this work, Mo2C with the two-dimensional ultrathin nanosheet structure on the surface and excellent conductivity was successfully employed as nanocarriers for the capture of ECL reagent of luminol-AuNPs (luminol-AuNPs@Mo2C). Notably, the luminol-AuNPs in the hybrids exhibited enhanced ECL performance (ca. 6-fold) as compared with the individual luminol-AuNPs due to the facilitated electron transfer process. Ultimately, the as-prepared ECL probe was used to construct a label-free ECL immunosensor. Here, α-fetoprotein (AFP) was chosen as a model because AFP has been widely considered as a tumor marker. The elevated concentration of AFP in adult plasma often indicated some cancerous diseases, such as hepatocellular cancer, endoderm carcinoma, testicular cancer, teratoma, yolk sac cancer, ovarian cancer and liver metastasis from gastric cancer.40-42 As illustrated in Scheme 1, the loaded luminol-AuNPs offered abundant binding sites for anchoring bio-recognized elements. With the increase of the AFP, the ECL signal decreased gradually due to the insulting property of protein molecules and the steric hindrance effect. Satisfied results with good linear relationship and practical application were obtained using the present sensing interface.

EXPERIMENTAL SECTION ACS Paragon Plus Environment

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Chemicals and Reagents Luminol was obtained from Sigma-Aldrich (Milwaukee, WI, USA). HAuCl4 was purchased from Shanghai Chemical Factory (Shanghai, China). Bovine serum albumin (BSA) and ammonium heptamolybdate tetrahydrate ((NH4)6Mo7O24·4H2O) were purchased from Sinopharm Chemical Reagent Co. Ltd. (Beijing, China). Dopamine and branched polyethyleneimine (BPEI, Mw = 10000) were obtained from Alfa Aesar (Ward Hill, MA, USA). NH3·H2O was purchased from Beijing Chemical Reagent (Beijing, China). Carcinoembryonic antigen (CEA), IgG, AFP and anti-AFP were obtained from Biocell Biotechnology Co. Ltd. (Zhengzhou, China). All of other reagents were of analytical grade. 0.1 M phosphate buffered saline (PBS) with the pH of 7.4 was used for incubation, while 0.2 M carbonate buffer solution (CBS) with different pH values was used for the optimization of ECL detection. 0.2 M CBS (pH 9.6) containing 5 mM H2O2 was chosen as the working buffer. All of the solutions were prepared by the deionized Millipore Mill-Q water (18.2 MΩ·cm-1). Apparatus. Transmission electron microscopy (TEM) images were obtained using an H-8100 EM transmission electron microscope at an accelerating voltage of 100 kV (Hitachi, Tokyo, Japan). Scanning electron microscope (SEM) images were performed on an XL30 ESEM FEG SEM (Philips, Netherlands). Ultraviolet-visible (UV-vis) absorption spectra were recorded on a Cary 500 Scan UV-vis spectrophotometer (Varian, Harbor City, CA, USA). The Zeta potential values were taken on a Zetasizer Nano-Z system (Malveren Instruments). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were collected on a CHI 660E electrochemical workstation (Chenhua, Shanghai, China). ECL were carried out on a model MPI-A capillary electrophoresis-ECL system (Xi’an Remex Electronics Co. Ltd., Xi’ an, China). Synthesis of luminol-AuNPs. luminol-AuNPs were prepared according to the previous work.19 In brief, 1.0 mL 0.01 M luminol solution prepared in 0.1 M NaOH was added in 50 mL boiling HAuCl4 solution (0.01%, w/w) immediately under stirring. The above solution was kept for 30 min at the boiling point and the color was changed from yellow to black, then to purple and a wine red at last in this process. After removing the heating source, the mixture continued stirring for another 20 min. Finally, the prepared ACS Paragon Plus Environment


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luminol-AuNPs were centrifuged and washed with distilled water for several times and then stored at 4 °C. Preparation of Mo2C/BPEI. The Mo2C nanotubes were obtained via a template-consuming strategy and high temperature annealing from MoO3 nanorods according to the previous study.39 Then 3 mL of 1 mg·mL-1 Mo2C was dispersed in 12 mL mixed solution of 0.625 M KCl and 1.25 mg·mL-1 BPEI. After sonication and shaking overnight, a homogeneous black solution was achieved. Finally, the precipitate was collected by centrifugation, and washed with water for several times before re-dispersed in 6 mL distilled water to remove the non-specific adsorption of BPEI. Construction of luminol-AuNPs@Mo2C nanocomposite. The luminol-AuNPs@Mo2C nanocomposite was prepared by adding Mo2C/BPEI into the luminol-AuNPs colloid dropwisely along with sonication and shaking vigorously. After reacting overnight, the composite was collected and washed by centrifugation at 7000 rpm. Preparation and Fabrication of the label-free ECL immunosensor. The glassy carbon electrode (GCE) was polished with 0.3 and 0.05 µm alumina slurry, followed by sonication in water, ethanol and water sequentially and drying under nitrogen. Then, 5 µL of the luminol-AuNPs@Mo2C suspension was dropped on the GCE, and after drying in the room temperature, 5 µL 100 µg·mL-1 anti-AFP was dropped onto it and incubated for 12 h at 4 °C to get the anti-AFP/luminol-AuNPs@Mo2C bioconjugate. In order to block the nonspecific binding site, 5 µL BSA (0.05 mg·mL-1) was added and incubated at room temperature for 1 h. After rinsing with 0.1 M PBS buffer (PH 7.4), the label-free ECL immunosensor was fabricated and stored at 4 °C for the recognization of AFP. ECL detection of AFP. Prior to detect, the label-free ECL sensing platform was incubated with different concentrations of AFP at 37 °C For 1 h in 0.1 M PBS buffer (pH 7.4). After washing with 0.1 M PBS buffer (pH 7.4), the prepared electrode was stored at 4 °C. The ECL behavior was carried out on a model MPI-A capillary electrophoresis-ECL system with a typical three-electrode system. A modified GCE was used as ACS Paragon Plus Environment

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working electrode, while Ag/AgCl with saturated KCl and Pt wire was used as the reference and counter electrodes, respectively. They were immersed into the electrolyte in an electrolytic cell with a transparent bottom. The ECL was carried out under pulsed potential condition with the initial potential of 0 V, pulse potential of 0.9 V, pulse period of 10 s and pulse time of 1 s in 0.2 M CBS buffer containing 5 mM H2O2, which was similar to our previous work.17 For proving the practical application, the ELISA experiments were employed (the sample was prepared using two-fold dilution and standard curve was drawn using AFP with different concentrations).

RESULT AND DISCUSSION

Characterization of the luminol-AuNPs@Mo2C nanocomposite. Firstly, the self-degraded template of the MoO3 nanorods was prepared via a facile hydrothermal method.43 The morphology and structure of the MoO3 nanorods were characterized by TEM, SEM, and X-ray diffraction (XRD). As shown in Fig. S1, the MoO3 nanorod with a very smooth surface has an average diameter of 200 nm and high crystallinity according to the XRD survey (Fig. S2A), which is consistent with the previous work.39 Then, MoO42- and dopamine hydrochloride were introduced into the MoO3 nanorod suspension as Mo and C precursor. In the presence of ammonia solution, MoO3 nanorods were dissolved and Mo-polydopamine nanotubes with hierarchical architecture were formed. After annealing at 750 °C for 10 h in Ar conditions, Mo2C nanotubes with hollow interior were formed, as confirmed from the cracked nanotube in the Figure 1B. Compared with the MoO3 nanorods, the walls of the Mo2C nanotubes are rough and composed of many two-dimensional ultrathin nanosheets structure (Figure 1B), which would increase the specific surface area significantly and offer more opportunities to catch the functional molecules for subsequent immobilization of luminol-AuNPs. XRD pattern of Mo2C nanotubes (Figure S2B) indicated that the most of the characteristic diffraction peaks could be readily indexed to the hexagonal phase Mo2C crystal structure (JCPDS card no. 35-0787). Luminol-AuNPs was synthesized based on the luminol as capping and reduced reagents, therefore luminol and 3-aminophthalate were co-existed and coadsorbed onto the surface of AuNPs.19 The morphology of the formed luminol-AuNPs was characterized by TEM. As depicted in Figure S3, the colloids were well-dispersed with a particle diameter about 20 nm. The surface plasmon absorption wavelength of the as-prepared luminol-AuNPs was centered at 520 nm, indicating the successful ACS Paragon Plus Environment


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formation of Au colloids. The luminol-AuNPs@Mo2C nanocomposites were constructed via a simple self-assembly procedure with BPEI as bridging molecule. As shown in Figure S4, the zeta potential increased to 36.2 mV after modification with BPEI and therefore large amount of negatively charged luminol-AuNPs were captured on the Mo2C nanotubes based on the electrostatic interaction and the covalent interaction between Au and N atoms44. From the TEM and SEM images of the luminol-AuNPs@Mo2C composites (Figure 1D and E), luminol-AuNPs were clearly observed on the surface of Mo2C-BPEI nanotubes, indicating the successful adhesion of luminol-AuNPs. To further obtain the information of loading amount of luminol-AuNPs, ICP-MS was introduced and about 40% of luminol-AuNPs was captured on the functionalized Mo2C via Au-N bonding and electrostatic interaction. Moreover, the element mapping images also indicated (Figure 1C and F) the uniform distribution of luminol-AuNPs on the surface of Mo2C, which provided more anchoring sites for the further modification of biomolecules. ECL and Electrochemical behavior of luminol-AuNPs@Mo2C. The ECL behavior of luminol-AuNPs loaded on the multifunctional nanocomposites was evaluated (Figure 2A). As we know, the ECL feature of luminol has a great dependence on the pH of the solution. Consequently, the effect of pH using 0.2 M CBS buffer was optimized in the range of 9.2 to 10.2 firstly and an optimal ECL signal was obtained at pH 9.6 (as Figure S5). As a control, the ECL performance of bare GCE, Mo2C and luminol-AuNPs were collected under the optimized condition. As expected, no ECL emission signals were observed on the bare GCE and Mo2C modified electrodes. Surprisingly, the ECL intensity with the luminol-AuNPs@Mo2C nanocomposites was obviously stronger (ca. 6-fold) than that of the luminol-AuNPs. In order to further understand the reason of the enhanced ECL efficiency, the electrochemical behavior of the luminol-AuNPs@Mo2C nanocomposites was investigated using CV and EIS in 100 mM KCl solution containing 5.0 mM [Fe(CN)6]3-/4-, and the results were demonstrated in Figure 2B and C. It can be obviously seen in CV curves (Figure 2B) that Mo2C modified GCE exhibited the highest redox peak current (curve a), which may be attributed to the excellent electrical conductivity and the high specific surface. For the luminol-AuNPs@Mo2C electrode, the redox peak decreased slightly due to the negative charged interface, which may repel the [Fe(CN)6]3-/4- anion and retard the electron transfer. Notably, compared with the bare GCE and luminol-AuNPs electrode, both Mo2C and ACS Paragon Plus Environment

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luminol-AuNPs@Mo2C electrode modified GCE exhibited the efficient electron transfer ability, indicating the possibility of Mo2C as conducting pathway for the construction of ultrasensitive ECL probe. Consistent results were also achieved with the EIS (Figure 2C) by monitoring the interface change of electron-transfer resistance (Ret). With Mo2C modified GCE, a very small semicircle domain was observed, indicating the fast response of electron transfer and lower Ret. Although the introduction of negative luminol-AuNPs onto the surface of the Mo2C nanocarriers led to an increment of Ret due to the electrostatic repulsion, the luminol-AuNPs@Mo2C modified electrode still manifested fast response relative to bare GCE and luminol-AuNPs electrode. All these results indicated that Mo2C as nanocarriers could accelerate the electron transfer efficiently owing to the 2D ultrathin nanosheet structure of the walls and the high specific surface area. It was determined that in the prepared probe the Mo2C was not only used as the prominent electronic conductivity to improve the electron transfer at the electrode, but also acted as the efficient nanocarriers to load luminol-AuNPs efficiently, thus improving the ECL efficiency. Feasibility of the ECL immunosensor in AFP detection. Given the excellent luminescence behavior, luminol-AuNPs@Mo2C nanocomposites were further used to construct a sensitive label-free ECL immunosensor for the detection of AFP. Figure 3A displays the CV characterization of the fabrication procedure of the immunosensor. When anti-AFP, BSA and AFP were assembled on the surface of luminol-AuNPs@Mo2C nanocomposite in succession; the peak currents decreased gradually indicating the successful construction. The assemble procedure on the surface of the luminol-AuNPs@Mo2C nanocomposite was also confirmed by the increase of the Ret. As depicted in Figure 3B, the electrode modified with luminol-AuNPs@Mo2C nanocomposite showed a very small resistance due to the high electronic conductivity of Mo2C nanotubes. After incubated with anti-AFP, a remarkable increment of Ret was observed since the antibody hindered the electron transfer. Subsequent blocking the residual nonspecific binding sites with BSA resulted in the further increase of the Ret, which may be due to the inhibitory effect of BSA as the inert electron layer45. With the further incubation by AFP, antigen-antibody complex was formed and an increment of Ret is observed, indicating the electron transfer kinetics of the redox probe was hindered by the insulating property and steric hindrance effect of antibody-antigen complex. To demonstrate the ECL feasibility of the immunosensor, the ECL performance was collected using ACS Paragon Plus Environment


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different functionalized electrodes. As shown in Figure 3C, after the immobilization of the antibody and BSA, an obvious ECL decrease was observed due to the insulting property of protein molecules and the steric hindrance effect. With the further incubation by AFP, the resistance of the AFP further inhibited interfacial electron transfer and decreased ECL was obtained, demonstrating the successful construction of the signal-off ECL sensor platform for AFP detection. Compared with the results of CV and EIS (Figure 3A), ECL sensing platform exhibited higher sensitivity which endows the sensor platform with better performance. Performance of the ECL immunosensor in AFP detection. The ECL immunosensor assay for highly specific and sensitive detection of AFP was also investigated. Under the optimized condition, the ECL immunosensor was used to detect AFP. Figure 4A shows the dependence of ECL response of the immunosensor after incubation on different concentration of AFP. It was obviously that the ECL intensity decreased gradually with the increase of AFP concentration, a good linear correlation between the ECL response and the logarithm of the concentration of the AFP in a range of 0.1 pg·mL-1 to 30 ng·mL-1 was obtained in the Figure 4. The linear regression equation was I = -3795 lg c + 6989 with a correlation coefficient of 0.998 (n=3), and the limit of detection was as low as 0.03 pg·mL-1 (S/N=3). The results indicate that the proposed ECL sensor is highly sensitive and holds a great potential in detecting AFP. Compared with other strategies for AFP detection shown in Table S1, this luminol-AuNPs@Mo2C based ECL immunosensor displays excellent analytical performance with wider detection range and lower LOD. Prominent stability is a crucial factor for the practical application of the ECL immunosensor. Figure 5A shows the ECL response of the immunosensor using 20 ng·mL-1 AFP under ten cycles of successive pulse potential, which was implemented in 0.2 M CBS buffer with pH of 9.6 containing 5 mM H2O2. It is obviously that a rather stable ECL emission has been achieved and the relative standard deviation (RSD) is 2.36%, which confirms that the proposed platform exhibited an excellent stability in the AFP detection and holds a promising potential for practical application. In order to assess the selectivity of the proposed ECL sensor, several interfering proteins such as CEA, BSA and IgG were chosen using the constructed ECL sensor as controls. The concentration of the interfering proteins was all 20 ng·mL-1, which was about 20 times higher than the concentration of AFP (1 ng·mL-1), and the mixture contained all of them. As shown in Figure 5B, the ECL response of the ACS Paragon Plus Environment

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interfering proteins didn’t show significant difference compared with the blank solution. Only the presence of AFP led to the obvious ECL decrease, which indicates that the as-prepared ECL platform has excellent selectivity toward the AFP. Analytical performance in real samples. The practical application of the ECL immunosensor was investigated by detecting AFP in human serum. The concentration of AFP in the diluted serum sample (0.39 ng·mL-1) determined with this method was consistent with ELISA (as shown in Figure S8). Moreover, the recovery experiment was also done by adding the standard AFP solution into the human serum using the standard curve method. The as-fabricated immunosensor exhibited excellent performance in AFP detection with the recovery of 99.55% to 101.4% (Table S2). CONCLUSION In summary, we have prepared a multifunctinalized nanocomposite based on the Mo2C as the ECL probe nanocarriers and used it for the fabrication of a simple and sensitive ECL sensing platform. The introduction of the unique structure of Mo2C with the two-dimensional nanosheet and excellent electron transfer ability combined with intrinsic ECL behavior of loaded luminol-AuNPs endowed the multifunctinalized nanocomposites stronger ECL emission (ca. 6-fold) compared with that obtained using the individual luminol-AuNPs. A label-free and solid-state ECL immunosensor was finally constructed with ultrasensitivity, good selectivity and excellent stability for the detection of AFP and successfully used in real human serum samples. Meanwhile, this novel sensing interface can be extended to other biomolecules or biomarker. ASSOCIATED CONTENT Supporting Information Additional information about the TEM image, SEM image, XRD spectra, UV-vis absorption spectra, Zeta potential values, the comparative table with the previous work, the table on results of the ECL in the blood sample and ELISA results. The Supporting Information is available free of charge on the ACS Publications website. AUTHOR INFORMATION ACS Paragon Plus Environment


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Corresponding Author *Tel: +86-431-85262003. Fax: +86-431-85689711. E-mail: [email protected]. *Tel: +86-431-85262003. Fax: +86-431-85689711. E-mail: [email protected]. Notes The authors declare no competing financial interest. ACKNOWLEDGMENT This work was supported by the National Natural Science Foundation of China (Grant No. 21427811), MOST China (No. 2016YFA0203200, 2016YFA0201300 and 2013YQ170585), Youth Innovation Promotion Association CAS (No.2016208) and Jilin Province Science and Technology Development Plan Project 20170101194JC.


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FIGURE CAPTIONS Scheme 1. Schematic illustration of the label-free immunosensor for AFP detection. Figure 1. TEM and SEM images of Mo2C (A, B) and luminol-AuNPs (D, E), element mapping images of luminol-AuNPs@Mo2C (C, F). Figure 2. ECL response (A) in 0.2 M CBS buffer (pH 9.6) containing 5.0 mM H2O2, CV (B) and EIS (C) characterization in 0.1 M KCl aqueous solution containing 5.0 mM [Fe(CN)6]3-/4- of the modification of GCE: Mo2C (a), luminol-AuNPs@Mo2C (b), bare GCE (c) and luminol-AuNPs (d). Figure 3. CV (A) and EIS (B) characterization in 0.1 M KCl aqueous solution containing 5.0 mM [Fe(CN)6]3-/4- and ECL response (C) in 0.2 M CBS buffer (pH 9.6) containing 5 mM H2O2 of the modification of GCE: luminol-AuNPs@Mo2C (a), luminol-AuNPs@Mo2C/anti-AFP(b), luminol-AuNPs@Mo2C/anti-AFP/BSA(c) and luminol-AuNPs@Mo2C/anti-AFP/BSA/AFP (d). Figure 4. (A) The ECL reponse of different concentrations of AFP in 0.2 M CBS buffer (pH 9.6) containing 5 mM H2O2: 0.0001 ng·mL-1 (a), 0.001 ng·mL-1 (b), 0.01 ng·mL-1 (c), 0.1 ng·mL-1 (d), 1 ng·mL-1 (e), 5 ng·mL-1 (f), 10 ng·mL-1 (g), 20 ng·mL-1 (h), 30 ng·mL-1 (i). (B) Calibration curve of AFP detection. Figure 5. (A) Stability of the ECL immunosensor incubated with 20 ng·mL1 AFP in 0.2 M CBS buffer (pH 9.6) containing 5 mM H2O2 under ten cycles of continues pulse potential. (B) Selectivity of the ECL sensor: blank, CEA (20 ng·mL-1), BSA (20 ng·mL-1), IgG (20 ng·mL-1), AFP (1 ng·mL-1) and Mixture (containing all the above analytes).



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Scheme 1.


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Figure 1.



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Figure 2.


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Figure 3.



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Figure 4.


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Figure 5.



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For TOC only


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High-sensitivity electrochemiluminescence probe ... - ACS Publications

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