Elsevier

Biosensors and Bioelectronics

Volume 94, 15 August 2017, Pages 141-147
Biosensors and Bioelectronics

Dual-responsive electrochemical immunosensor for prostate specific antigen detection based on Au-CoS/graphene and CeO2/ionic liquids doped with carboxymethyl chitosan complex

https://doi.org/10.1016/j.bios.2017.03.001Get rights and content

Highlights

  • DPV and amperometric i–t curve methods were combined to test the electrochemical immunosensor for the first time.

  • Amplified amperometric i–t signals can be generated from Au-CoS/graphene modified electrode.

  • Amplified DPV signals can be generated from label of TB/M-CeO2/CMC/ILs.

  • CMC/ILs prevent the leak of TB and facilitate the electron transfer of NH2-M-CeO2.

Abstract

Combining dual-signal channels of differential pulse voltammetry (DPV) and amperometric i–t curve, we developed an amplified sandwich-type electrochemical immunosensor for ultrasensitive detection of prostate specific antigen (PSA). Due to the large specific surface area and good adsorption property, 3-aminopropyltriethox functionalized CeO2 mesoporous nanoparticles (NH2-M-CeO2) was used for supporting toluidine blue (TB) as the electron transfer mediator and anti-PSA as the signal response. To prevent the leak of TB and facilitate the electron transfer property of NH2-M-CeO2, ionic liquids doped carboxymethyl chitosan (CMC/ILs) were incorporated and TB/M-CeO2/CMC/ILs complex was gotten to act as the label of anti-PSA (Ab2). Meanwhile, Au-CoS/graphene was as the matrix materials modifying the electrode to immobilize anti-PSA (Ab1). Specifically, Au-CoS/graphene cannot produce electrochemical signals through DPV method but can provide an obviously electrochemical signal towards catalysis hydrogen peroxide (H2O2) through amperometric i–t curve method. At the time of detection, with the increase of concentrations of PSA, the DPV signals from TB/M-CeO2/CMC/ILs were increased and the amperometric i–t curve signals from the Au-CoS/graphene were decreased. Under the optimum conditions, the immunosensor demonstrated remarkable analytical performance of a linear range of 0.5 pg/mL to 50 ng/mL with a detection limit of 0.16 pg/mL for quantitative detection of PSA (S/N=3).

Introduction

Prostate cancer is one of most common malignant tumor that the mortality rate ranks second in various kinds of cancer amongst men (Center et al., 2012; Çevik et al., 2016). The early detection of tumor marker is one of great effective methods for early identification and diagnosis cancer (Wang et al., 2015b). Prostate specific antigen (PSA), as a reliable tumor marker, the concentration is directly correlated with the chance of prostate cancer (Moon et al., 2014). In general, PSA levels in 4.0–10.0 ng/mL indicate a 25–40% probability of prostate cancer, the cancer risk significantly increases to 67% if the PSA level is greater than 10 ng/mL (Leibovici et al., 2005). However, lower PSA levels (<4.0 ng/mL) still have been associated with a high incidence of prostate cancer during the past few years (Yang et al., 2006). Therefore, the scope of limit of detection in the nano-gram to femto-gram is greatly necessary (Gretzre et al., 2002).

Currently, using various detection signals based on electrochemistry, optics, mass, or target analytes in the human serum specimens could realize early identification and detection even at very low levels (Elrifai et al., 2016, Lee et al., 2013, Li et al., 2016a, Li et al., 2016b). Among these promising analytical approaches, electrochemical detection has attracted much interest because it allows direct, specific, and real-time monitoring (Bai et al., 2012, Jayanthi et al., 2017, Li et al., 2012). The dual-signal property of the electrochemical immunosensor has readily increased analytical performance and clinical reliability, thus improved the accuracy of the results. However, combining dual-signal channels of DPV and amperometric i–t curve have not used for sensitive detection of tumor markers.

CeO2 mesoporous nanoparticle (M-CeO2) has been massively applied to load other nano-materials, chemicals or biomolecules because of its good adsorption property, high surface area, uniform size distribution, well-defined pore topology and good water dispersibility (Liang et al., 2010). Toluidine blue (TB), a widely used electron transfer mediator, is loaded on M-CeO2 to produce electrochemical signals through physical absorption and cross-link effect of 1-ethyl-3-(3-dimethyla-minopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) (Yao and Shiu, 2007, Li et al., 2016a, Li et al., 2016b). In addition, ionic liquids doped carboxymethyl chitosan (CMC/ILs) can offer enough adsorption groups (–OH, –COOH and –NH–) to increase adsorption capacity towards TB. On the other side, CMC/ILs also prevents TB from leaking and effectively facilitates the electron transfer (Lemraski and Kargar, 2014). Using gold nano-particles functionalized CoS/graphene sheets composite (Au-CoS/graphene) as the matrix materials which cannot produce electrochemical signals through differential pulse voltammetry (DPV) method, but it has good electrical conductivity to contribute to improve the sensitivity in this designed immunosensor.

L-Cysteine (HSCH2CHNH2COOH), an environmental-friendly small bio-molecule, has been employed to prepare various metal sulfides because it contains multifunctional groups in molecule (–SH, –NH2 and –COOH) (Qiu et al., 2014, Wen et al., 2009). These groups not only make L-cysteine as a kind of commonly used self-assembly reagent in the preparation of various metal sulfides nanomaterials, but also are employed as reductant to reduce graphene oxide sheets under hydrothermal condition (Chen et al., 2015). Graphene sheets have been widely applied in the field of immunosensor due to their high surface area-to-volume ratio, good electron transfer property (Ge et al., 2014, Wang et al., 2015a, Zhang et al., 2017, Zhu et al., 2017). In this work, L-cysteine has been employed to prepare CoS particles and they with good catalytic performance effectively catalyze the reduction of hydrogen peroxide (H2O2), but these CoS particles have agglomerated together due to minimizing the surface energy. Therefore, CoS particles as the matrix materials which are not conducive to immobilize Ab1. In contrast, the introduction of graphene oxide sheets which can not only make CoS particles smaller, but also effectively prevent the CoS particles from agglomerating. Because between the CoS particles and graphene sheets has good synergistic coupling effects, Au-CoS/graphene composite not only provides an obviously electrochemical signal towards catalysis H2O2 through amperometric i–t curve method, but also immobilizes abundant primary anti-PSA (Ab1) due to stable conjunction between noble metal nanoparticles and amino groups and further promotes electron transfer. NH2-M-CeO2 possesses the large steric hindrance and bad electrocatalytic activity to further improve the sensitivity of the immunosensor through amperometric i–t curve method. As a result, the DPV signal from TB/M-CeO2/CMC/ILs increased, and the amperometric i–t curve signal from the Au-CoS/graphene decreased correspondingly with the increase of concentrations of PSA.

Therefore, the dual-signal property of the electrochemical immunosensor is responsible for an interaction with Au-CoS/graphene and TB/M-CeO2/CMC/ILs complex in the detection of tumor markers on the basis of two different electrochemical signals change, readily increasing analytical performance and clinical reliability, thus improving the accuracy of the results.

Section snippets

Reagents and apparatuses

Human antibody-PSA and antigen-PSA were bought from Biocell Science Co., Ltd. (Shanghai, China). Bovine serum albumin (BSA, ≥99%) was purchased from Sigma Reagent Co., Ltd. (St. Louis, MO, USA), 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC, 98.5%), N-hydroxysuccinimide (NHS, 98%) and 3-amino- propyltriethoxysilane (APTES, 98%) were purchased Shanghai Aladdin (Chemistry Co., Ltd., China). 1-butyl-pyridine tetrafluoroborate (>99%) was purchased from Lanzhou Institute of Chemical Physics.

Characterization of materials

Fig. 2(A, B) illustrates the SEM images of the pristine CoS particles and CoS/graphene composite. As revealed in Fig. 2(A), the pristine CoS displays irregular particles, the most of which ranges from 60 nm to 70 nm in size. It is also noteworthy that CoS particles have agglomerated together to minimize the surface energy. In contrast, along with the introduction of graphene oxide sheets, the CoS particles exhibited much smaller and more uniform sizes than the pristine CoS particles as presented

Conclusions

This work has developed a dual-responsive electrochemical immunosensor (DPV and amperometric i–t curve) for quantitative detection of PSA based on Au-CoS/graphene as the matrix materials and TB/M-CeO2/CMC/ILs complex as the labels. M-CeO2 had adsorbed the electron transfer mediator TB to produce directly electrochemical signals. Ionic liquids (ILs) doped carboxymethyl chitosan (CMC) was employed to prevent the leak of TB and facilitate the electron transfer. Au-CoS/graphene as the matrix

Acknowledgments

This study was supported by the National Natural Science Foundation of China (Nos. 21375047, 21377046, 21575050 and 21505051), the Science and Technology Plan Project of Jinan (No. 201307010), the Science and Technology Development Plan of Shandong Province (No. 2014GSF120004), the Special Project for Independent Innovation and Achievements Transformation of Shandong Province (No. 2014ZZCX05101), Doctoral and Postdoctoral Science Foundation of University of Jinan. Qin Wei thanks the Special

References (29)

  • L. Bai et al.

    Biomaterials

    (2012)
  • M.M. Center et al.

    Eur. Urol.

    (2012)
  • E. Çevik et al.

    Biosens. Bioelectron.

    (2016)
  • Q. Chen et al.

    J. Power Sources

    (2015)
  • R. Feng et al.

    Biosens. Bioelectron.

    (2013)
  • A. Elrifai et al.

    Pol. Ann. Med.

    (2016)
  • S. Ge et al.

    Sens. Actuators B: Chem.

    (2014)
  • V.S. Jayanthi et al.

    Biosens. Bioelectron.

    (2017)
  • S.-W. Lee et al.

    Int. J. Adhes. Adhes.

    (2013)
  • D. Leibovici et al.

    Urol. Oncol.

    (2005)
  • E.G. Lemraski et al.

    J. Mol. Liq.

    (2014)
  • J. Li et al.

    Biosens. Bioelectron.

    (2016)
  • J.-M. Moon et al.

    Biosens. Bioelectron.

    (2014)
  • S. Peng et al.

    Nano Energy

    (2015)
  • Cited by (68)

    • CoNi-RGO and NiCo<inf>2</inf>S<inf>4</inf>-ZIF/g-C<inf>3</inf>N<inf>4</inf> signal amplified electrochemical immunosensors for sensitive detection of CYFRA 21-1

      2022, Analytical Biochemistry
      Citation Excerpt :

      Therefore, the development of signal amplification immunosensors is indispensable, which is conducive to the new height of electrochemical immunoassay biomarkers. It is recognized that transition metal sulfides, especially ternary transition metal sulfides, perform very well in the research of electrochemical immunosensors [21–23]. Transition metal sulfides have high stability, excellent conductivity and outstanding

    View all citing articles on Scopus
    View full text