Microchimica Acta

, 186:416 | Cite as

An electrochemical sandwich immunosensor for cardiac troponin I by using nitrogen/sulfur co-doped graphene oxide modified with Au@Ag nanocubes as amplifiers

  • Hui Lv
  • Xiaobo Zhang
  • Yueyun LiEmail author
  • Yong Ren
  • Chunyan Zhang
  • Ping Wang
  • Zhen Xu
  • Xinjin LiEmail author
  • Zhiwei Chen
  • Yunhui Dong
Original Paper


A voltammetric sandwich immunoassay is described for the biomarker cardiac troponin I (cTnI). The gold nanocube-functionalized graphene oxide (AuNC/GO) is employed as a substrate to accelerate the electron transfer and to immobilize more primary antibodies. It also employs composite materials prepared from bimetallic gold/silver core-shell nanocubes and nitrogen and sulfur co-doped reduced graphene oxide as the signal amplifier. The introduction of N and S into GO enlarges the active surface and accelerates the electron transfer rate. Such unique characteristics render the material an effective support substrate to load more Au@AgNC and to immobilize an increasing number of second antibodies via Ag-N bonds. After specific binding with cTnI, the immunosensor was incubated in a labeled cTnI secondary antibody solution. The amperometric signal change is then measured at 0.34 V (vs. SCE) using o-phenylenediamine and hydrogen peroxide as an electrochemical probe. Response is linear in the concentration range from 100 fg∙mL−1 to 250 ng∙mL−1, and the detection limit is 33 fg∙mL−1.

Graphical abstract

Schematic presentation of cardiac troponin I (cTnI) electrochemical immunosensor based on gold nanocube-functionalized graphene oxide (AuNC/GO) as substrate material, bimetallic gold/silver core-shell nanocubes and nitrogen and sulfur co-doped reduced graphene oxide (Au@AgNC/N, S-rGO) as signal amplifier, and hydrogen peroxide (H2O2) and o-phenylenediamine (o-PD) as redox probe.


Sandwich immunoassay Core-shell nanocube Multiple signal amplification Specific binding Electrochemical probe 



This study was financially supported by the Key Research and Development Program of Shandong Province (No. 2018GSF120001, 2018GNC110038), the National Natural Science Foundation of China (Nos. 21575079). All of the authors express their deep thanks.

Compliance with ethical standards

The author(s) declare that they have no competing interests.

All experiments were performed in compliance with relevant laws or guidelines of Shandong University of Technology and approved by the ethics committee at Shandong University of Technology, China. Moreover, informed consent was obtained from human participants of this study.

Supplementary material

604_2019_3526_MOESM1_ESM.docx (1.3 mb)
ESM 1 (DOCX 1.33 mb)


  1. 1.
    Sheng Q, Qiao X, Zhou M, Zheng J (2017) Recent progress in electrochemical sensing of cardiac troponin by using nanomaterial-induced signal amplification. Microchim Acta 184(6):1573–1585. CrossRefGoogle Scholar
  2. 2.
    Rezaei B, Shoushtari AM, Rabiee M, Uzun L, Mak WC, Turner APF (2018) An electrochemical immunosensor for cardiac troponin I using electrospun carboxylated multi-walled carbon nanotube -whiskered nanofibres. Talanta 182:178–176. CrossRefPubMedGoogle Scholar
  3. 3.
    Gao F, Fan T, Ou S, Wu J, Zhang X, Luo J, Li N, Yao Y, Mou Y, Liao X (2017) Highly efficient electrochemical sensing platform for sensitive detection DNA methylation, and methyltransferase activity based on Ag NPs decorated carbon nanocube. Biosens Bioelectron 99:201–208. CrossRefPubMedGoogle Scholar
  4. 4.
    Hong Q, Yang L, Ge L, Liu Z, Li F (2018) Direct-laser-writing of three-dimensional porous graphene frameworks on indium-tin oxide for sensitive electrochemical biosensing. Analyst 142(5):780–786. CrossRefGoogle Scholar
  5. 5.
    Wei Z, Guo S, Asaka K (2014) An insight into the capacitive properties of reduced graphene oxide. ACS Appl Mater Interfaces 6(4):2248–2254. CrossRefGoogle Scholar
  6. 6.
    Singh S, Tuteja SK, Sillu D, Deep A, Suri CR (2016) Gold nanoparticles-reduced graphene oxide based electrochemical immunosensor for the cardiac biomarker myoglobin. Microchim Acta 183(5):1729–1738. CrossRefGoogle Scholar
  7. 7.
    Lv H, Li Y, Zhang X, Gao Z, Zhang C, Zhang S, Dong Y (2018) Enhanced peroxidase-like properties of Au@Pt DNs/NG/Cu2+ and application of sandwich-type electrochemical immunosensor for highly sensitive detection of CEA. Biosens Bioelectron 112:1–7. CrossRefPubMedGoogle Scholar
  8. 8.
    Chen B, Jiang Z, Zhou L, Deng B, Jiang ZJ, Huang J, Liu M (2018) Electronic coupling induced high performance of N, S-codoped graphene supported CoS2 nanoparticles for catalytic reduction and evolution of oxygen. J Power Sources 389:178–187. CrossRefGoogle Scholar
  9. 9.
    Huo J, Zheng P, Wang X, Guo S (2018) Three-dimensional Sulphur/nitrogen co-doped reduced graphene oxide as high-performance supercapacitor binder-free electrodes. Appl Surf Sci 442:575–580. CrossRefGoogle Scholar
  10. 10.
    Jiang L, Han J, Li F, Gao J, Li Y, Dong Y, Wei Q (2015) A sandwich-type electrochemical immunosensor based on multiple signal amplification for α-fetoprotein labeled by platinum hybrid multiwalled carbon nanotubes adhered copper oxide. Electrochim Acta 160:7–14. CrossRefGoogle Scholar
  11. 11.
    Zhang X, Lv H, Li Y, Zhang C, Wang P, Liu Q, Ai B, Xu Z, Zhao Z (2019) Ultrasensitive sandwich-type immunosensor for cardiac troponin I based on enhanced electrocatalytic reduction of H2O2 by β-cyclodextrins functionalized 3D porous graphene supported Pd@Au nanocubes. J Mater Chem B 7(9):1460–1468. CrossRefGoogle Scholar
  12. 12.
    Zhang Y, Shuai Z, Zhou H, Luo Z, Liu B, Zhang Y, Zhang L, Chen S, Chao J, Weng L (2018) Single-molecule analysis of MicroRNA and logic operations using a smart plasmonic nanobiosensor. J Am Chem Soc 140(11):3988–3993. CrossRefPubMedGoogle Scholar
  13. 13.
    Zhang X, Li Y, Lv H, Feng J, Gao Z, Wang P, Dong Y, Liu Q, Zhao Z (2018) Sandwich-type electrochemical immunosensor based on Au@Ag supported on functionalized phenolic resin microporous carbon spheres for ultrasensitive analysis of α-fetoprotein. Biosens Bioelectron 106:142–148. CrossRefPubMedGoogle Scholar
  14. 14.
    Lin T, Zhang M, Xu F, Wang X, Xu Z, Guo L (2018) Colorimetric detection of benzoyl peroxide based on the etching of silver nanoshells of Au@Ag nanorods. Sensors Actuators B Chem 261:379–384. CrossRefGoogle Scholar
  15. 15.
    Xu H, Song P, Fernandez C, Wang J, Zhu M, Shiraishi Y, Du Y (2018) Sophisticated construction of binary PdPb alloy Nanocube as robust Electrocatalysts toward ethylene glycol and glycerol oxidation. ACS Appl Mater Interfaces 10:12659–12665. CrossRefPubMedGoogle Scholar
  16. 16.
    Chen P, Wang T, Zheng X, Tian D, Xia F, Zhou C (2018) An ultrasensitive electrochemical immunosensor based on C60-modified polyamidoamine dendrimer and Au NPs for co-catalytic silver deposition. New J Chem 42(6):4653–4660. CrossRefGoogle Scholar
  17. 17.
    Wang P, Li M, Pei F, Li Y, Liu Q, Dong Y, Chu Q, Zhu H (2017) An ultrasensitive sandwich-type electrochemical immunosensor based on the signal amplification system of double-deck gold film and thionine unite with platinum nanowire inlaid globular SBA-15 microsphere. Biosens Bioelectron 91:424–430. CrossRefPubMedGoogle Scholar
  18. 18.
    Li M, Wang P, Pei F, Yu H, Dong Y, Li Y, Liu Q, Chen P (2018) A novel signal amplification system fabricated immunosensor based on Au nanoparticles and mesoporous trimetallic PdPtCu nanospheres for sensitive detection of prostate specific antigen. Sensors Actuators B Chem 261:22–30. CrossRefGoogle Scholar
  19. 19.
    Fan F-R, Liu D-Y, Wu Y-F, Duan S, Xie Z-X, Jiang Z-Y, Tian Z-Q (2008) Epitaxial growth of heterogeneous metal nanocrystals: from gold nano-octahedra to palladium and silver nanocube. J Am Chem Soc 130(22):6949–6951. CrossRefPubMedGoogle Scholar
  20. 20.
    Zhang R, Zhang C, Zheng F, Li X, Sun C-L, Chen W (2018) Nitrogen and sulfur co-doped graphene nanoribbons: a novel metal-free catalyst for high performance electrochemical detection of 2, 4, 6-trinitrotoluene (TNT). Carbon 126:328–337. CrossRefGoogle Scholar
  21. 21.
    Marcano DC, Kosynkin DV, Berlin JM, Sinitskii A, Sun Z, Slesarev A, Alemany LB, Lu W, Tour JM (2010) Improved synthesis of graphene oxide. ACS Nano 4(8):4806–4814. CrossRefPubMedGoogle Scholar
  22. 22.
    Chen J, Kong L, Sun X, Feng J, Chen Z, Fan D, Wei Q (2018) Ultrasensitive photoelectrochemical immunosensor of cardiac troponin I detection based on dual inhibition effect of Ag@Cu2O core-shell submicron-particles on CdS QDs sensitized TiO2 nanosheets. Biosens Bioelectron 117:340–346. CrossRefPubMedGoogle Scholar
  23. 23.
    Sandil D, Srivastava S, Malhotra B, Sharma S, Puri NK (2018) Biofunctionalized tungsten trioxide-reduced graphene oxide nanocomposites for sensitive electrochemical immunosensing of cardiac biomarker. J Alloys Compd 763:102–110. CrossRefGoogle Scholar
  24. 24.
    Jiang M-H, Lu P, Lei Y-M, Chai Y-Q, Yuan R, Zhuo Y (2018) Self-accelerated electrochemiluminescence emitters of Ag@SnO2 nanoflowers for sensitive detection of cardiac troponin T. Electrochim Acta 271:464–471. CrossRefGoogle Scholar
  25. 25.
    Kazemi SH, Ghodsi E, Abdollahi S, Nadri S (2016) Porous graphene oxide nanostructure as an excellent scaffold for label-free electrochemical biosensor: detection of cardiac troponin I. Mater Sci Eng C Mater Biol Appl 69:447–452. CrossRefPubMedGoogle Scholar
  26. 26.
    Liu G, Meng Q, Zhang Y, Cao C, Goldys EM (2016) Nanocomposites of gold nanoparticles and graphene oxide towards an stable label-free electrochemical immunosensor for detection of cardiac marker troponin-I. Anal Chim Acta 909:1–8. CrossRefPubMedGoogle Scholar
  27. 27.
    Yan H, Tang X, Zhu X, Zeng Y, Lu X, Yin Z, Lu Y, Yang Y, Li L (2018) Sandwich-type electrochemical immunosensor for highly sensitive determination of cardiac troponin I using carboxyl-terminated ionic liquid and helical carbon nanotube composite as platform and ferrocenecarboxylic acid as signal label. Sensors Actuators B Chem 277:234–240. CrossRefGoogle Scholar
  28. 28.
    Chekin F, Vasilescu A, Jijie R, Singh SK, Kurungot S, Iancu M, Badea G, Boukherroub R, Szunerits S (2018) Sensitive electrochemical detection of cardiac troponin I in serum and saliva by nitrogen-doped porous reduced graphene oxide electrode. Sensors Actuators B Chem 262:180–187. CrossRefGoogle Scholar
  29. 29.
    Singal S, Srivastava AK, Gahtori B, Rajesh (2016) Immunoassay for troponin I using a glassy carbon electrode modified with a hybrid film consisting of graphene and multiwalled carbon nanotubes and decorated with platinum nanoparticles. Microchim Acta 183(4):1–10. CrossRefGoogle Scholar
  30. 30.
    Singal S, Srivastava AK, Dhakate S, Biradar AM, Rajesh R (2015) Electroactive graphene-multi-walled carbon nanotube hybrid supported impedimetric immunosensor for the detection of human cardiac troponin-I. RSC Adv 5(92):74994–75003. CrossRefGoogle Scholar
  31. 31.
    Zhang T, Ma N, Ali A, Wei Q, Wu D, Ren X (2018) Electrochemical ultrasensitive detection of cardiac troponin I using covalent organic frameworks for signal amplification. Biosens Bioelectron 119:176–181. CrossRefPubMedGoogle Scholar
  32. 32.
    Saxberg BE, Kowalski B (1979) Generalized standard addition method. Anal Chem 51(7):1031–1038. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Austria, part of Springer Nature 2019

Authors and Affiliations

  • Hui Lv
    • 1
  • Xiaobo Zhang
    • 1
  • Yueyun Li
    • 1
    Email author
  • Yong Ren
    • 2
  • Chunyan Zhang
    • 1
  • Ping Wang
    • 1
  • Zhen Xu
    • 1
  • Xinjin Li
    • 1
    Email author
  • Zhiwei Chen
    • 3
  • Yunhui Dong
    • 1
  1. 1.School of Chemistry and Chemical EngineeringShandong University of TechnologyZiboPeople’s Republic of China
  2. 2.Department of Mathematical SciencesZibo Normal CollegeZiboPeople’s Republic of China
  3. 3.School of Life ScienceShandong University of TechnologyZiboPeople’s Republic of China

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