Microchimica Acta

, 186:472 | Cite as

Disposable paper-based electrochemical sensor using thiol-terminated poly(2-methacryloyloxyethyl phosphorylcholine) for the label-free detection of C-reactive protein

  • Chanika Pinyorospathum
  • Sudkate Chaiyo
  • Pornpen Sae-ung
  • Voravee P. Hoven
  • Panittha Damsongsang
  • Weena Siangproh
  • Orawon ChailapakulEmail author
Original Paper


A paper-based electrochemical sensor is described that is based on the use of thiol-terminated poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC-SH) that was self-assembled on a gold nanoparticle-modified screen-printed electrode (SPE). The SPE sensor was used for label-free detection of C-reactive protein (CRP). Gold nanoparticles (AuNPs) were first electrodeposited on the SPCE, followed by the self-assembly of PMPC-SH on gold. The electrochemical response of the modified SPE to CRP was measured by differential pulse voltammetry (DPV). If the CRP on the paper device is contacted with Ca (II) ions, the current (measured by using hexacyanoferrate as the electrochemical probe) decreases. The signal drops in the 5 to 5000 ng·mL−1 CRP concentration range, and the lower detection limit (at 3 SD/slope) is 1.6 ng·mL−1. The use of a PMPC-modified surface also reduces the nonspecific adsorption of proteins. The sensor is not interfered by bilirubin, myoglobin and albumin. It was successfully applied to CRP detection in certified human serum. This sensor is applicable as an attractive protocol for an inexpensive, highly sensitive, and disposable material for electrochemical detection of CRP.

Graphical abstract

Schematic presentation of highly sensitive and disposable paper-based electrochemical sensor using thiol-terminated poly(2-methacryloyloxyethyl phosphorylcholine) in the presence of Ca2+ for the label-free C-reactive protein detection. The current was measured by differential pulse voltammetry.


C-reactive protein Paper-based analytical devices Differential pulse voltammetry Gold nanoparticles Phosphorylcholine 



CP thankfully acknowledges financial support from Thailand Research Fund, through the Royal Golden Jubilee Ph.D. Program (Grant No. PHD/0032/2558). This research was financially supported by the Thailand Research Fund through Research Team Promotion Grant (RTA6080002). This research was partially supported by Ratchadapiseksomphot Endowment Fund under Outstanding Research Performance Program (SciSuperIII), the National Nanotechnology Center (NANOTEC), NSTDA, Ministry of Science and Technology, Thailand, through its program of Research Network NANOTEC (RNN), and the Thailand Research Fund (RSA5980071).

Compliance with ethical standards

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

Supplementary material

604_2019_3559_MOESM1_ESM.docx (2.1 mb)
ESM 1 (DOCX 2192 kb)


  1. 1.
    Pepys MB, Hirschfield GM (2003) C-reactive protein: a critical update. J Clin Invest 111(12):1805–1812. CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Steel DM, Whitehead AS (1994) The major acute phase reactants: C-reactive protein, serum amyloid P component and serum amyloid a protein. Immunol Today 15(2):81–88. CrossRefPubMedGoogle Scholar
  3. 3.
    Buckley DI, Fu R, Freeman M, Rogers K, Helfand M (2009) C-reactive protein as a risk factor for coronary heart disease: a systematic review and meta-analyses for the u.s. preventive services task force. Ann Intern Med 151(7):483–495. CrossRefPubMedGoogle Scholar
  4. 4.
    Pepys MB, Hirschfield GM, Tennent GA, Ruth Gallimore J, Kahan MC, Bellotti V, Hawkins PN, Myers RM, Smith MD, Polara A, Cobb AJA, Ley SV, Andrew Aquilina J, Robinson CV, Sharif I, Gray GA, Sabin CA, Jenvey MC, Kolstoe SE, Thompson D, Wood SP (2006) Targeting C-reactive protein for the treatment of cardiovascular disease. Nature 440:1217–1221. CrossRefPubMedGoogle Scholar
  5. 5.
    Shrivastava AK, Singh HV, Raizada A, Singh SK (2015) C-reactive protein, inflammation and coronary heart disease. The Egyptian Heart Journal 67(2):89–97. CrossRefGoogle Scholar
  6. 6.
    Casas JP, Shah T, Hingorani AD, Danesh J, Pepys MB (2008) C-reactive protein and coronary heart disease: a critical review. J Intern Med 264(4):295–314. CrossRefPubMedGoogle Scholar
  7. 7.
    Şişman AR, Küme T, Taş G, Pn A, Pn T (2007) Comparison and evaluation of two C-reactive protein assays based on particle-enhanced immunoturbidimetry. J Clin Lab Anal 21(2):71–76. CrossRefPubMedGoogle Scholar
  8. 8.
    Chikkaveeraiah BV, Bhirde AA, Morgan NY, Eden HS, Chen X (2012) Electrochemical Immunosensors for detection of Cancer protein biomarkers. ACS Nano 6(8):6546–6561. CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Albareda-Sirvent M, Merkoçi A, Alegret S (2000) Configurations used in the design of screen-printed enzymatic biosensors. A review. Sensors Actuators B Chem 69(1):153–163. CrossRefGoogle Scholar
  10. 10.
    Hernández-Santos D, González-García MB, García AC (2002) Metal-Nanoparticles Based Electroanalysis. Electroanalysis 14(18):1225–1235.<1225::AID-ELAN1225>3.0.CO;2-Z CrossRefGoogle Scholar
  11. 11.
    Agüí L, Peña-Farfal C, Yáñez-Sedeño P, Pingarrón JM (2007) Electrochemical determination of homocysteine at a gold nanoparticle-modified electrode. Talanta 74(3):412–420. CrossRefPubMedGoogle Scholar
  12. 12.
    Rattanarat P, Dungchai W, Cate D, Volckens J, Chailapakul O, Henry CS (2014) Multilayer paper-based device for colorimetric and electrochemical quantification of metals. Anal Chem 86(7):3555–3562. CrossRefPubMedGoogle Scholar
  13. 13.
    Rattanarat P, Dungchai W, Cate DM, Siangproh W, Volckens J, Chailapakul O, Henry CS (2013) A microfluidic paper-based analytical device for rapid quantification of particulate chromium. Anal Chim Acta 800:50–55. CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Panraksa Y, Siangproh W, Khampieng T, Chailapakul O, Apilux A (2018) Paper-based amperometric sensor for determination of acetylcholinesterase using screen-printed graphene electrode. Talanta 178:1017–1023. CrossRefPubMedGoogle Scholar
  15. 15.
    Ruecha N, Rangkupan R, Rodthongkum N, Chailapakul O (2014) Novel paper-based cholesterol biosensor using graphene/polyvinylpyrrolidone/polyaniline nanocomposite. Biosens Bioelectron 52:13–19. CrossRefPubMedGoogle Scholar
  16. 16.
    Thompson D, Pepys MB, Wood SP (1999) The physiological structure of human C-reactive protein and its complex with phosphocholine. Structure 7(2):169–177. CrossRefPubMedGoogle Scholar
  17. 17.
    Volanakis JE, Wirtz KWA (1979) Interaction of C-reactive protein with artificial phosphatidylcholine bilayers. Nature 281:155–157. CrossRefPubMedGoogle Scholar
  18. 18.
    Wu J-G, Wei S-C, Chen Y, Chen J-H, Luo S-C (2018) Critical study of the recognition between C-reactive protein and surface-immobilized Phosphorylcholine by quartz crystal microbalance with dissipation. Langmuir 34(3):943–951. CrossRefPubMedGoogle Scholar
  19. 19.
    Goda T, Kjall P, Ishihara K, Richter-Dahlfors A, Miyahara Y (2014) Biomimetic interfaces reveal activation dynamics of C-reactive protein in local microenvironments. Adv Healthc Mater 3(11):1733–1738. CrossRefPubMedGoogle Scholar
  20. 20.
    Shimada T, Yasui T, Yokoyama A, Goda T, Hara M, Yanagida T, Kaji N, Kanai M, Nagashima K, Miyahara Y, Kawai T, Baba Y (2018) Biomolecular recognition on nanowire surfaces modified by the self-assembled monolayer. Lab Chip 18(21):3225–3229. CrossRefPubMedGoogle Scholar
  21. 21.
    Goda T, Toya M, Matsumoto A, Miyahara Y (2015) Poly(3,4-ethylenedioxythiophene) bearing Phosphorylcholine groups for metal-free, antibody-free, and low-impedance biosensors specific for C-reactive protein. ACS Appl Mater Interfaces 7(49):27440–27448. CrossRefPubMedGoogle Scholar
  22. 22.
    Kitayama Y, Takeuchi T (2014) Localized surface Plasmon resonance Nanosensing of C-reactive protein with poly(2-methacryloyloxyethyl phosphorylcholine)-grafted gold nanoparticles prepared by surface-initiated atom transfer radical polymerization. Anal Chem 86(11):5587–5594. CrossRefPubMedGoogle Scholar
  23. 23.
    Iwasaki S, Kawasaki H, Iwasaki Y (2019) Label-free specific detection and collection of C-reactive protein using Zwitterionic Phosphorylcholine-polymer-protected magnetic nanoparticles. Langmuir 35(5):1749–1755. CrossRefPubMedGoogle Scholar
  24. 24.
    Bhuchar N, Deng Z, Ishihara K, Narain R (2011) Detailed study of the reversible addition–fragmentation chain transfer polymerization and co-polymerization of 2-methacryloyloxyethyl phosphorylcholine. Polym Chem 2(3):632–639. CrossRefGoogle Scholar
  25. 25.
    Ma Q, Zhang H, Zhao J, Gong Y-K (2012) Fabrication of cell outer membrane mimetic polymer brush on polysulfone surface via RAFT technique. Appl Surf Sci 258(24):9711–9717. CrossRefGoogle Scholar
  26. 26.
    Jampasa S, Siangproh W, Duangmal K, Chailapakul O (2016) Electrochemically reduced graphene oxide-modified screen-printed carbon electrodes for a simple and highly sensitive electrochemical detection of synthetic colorants in beverages. Talanta 160:113–124. CrossRefPubMedGoogle Scholar
  27. 27.
    Katz E, Willner I (2003) Probing biomolecular interactions at conductive and Semiconductive surfaces by impedance spectroscopy: routes to Impedimetric Immunosensors, DNA-sensors, and enzyme biosensors. Electroanalysis 15(11):913–947. CrossRefGoogle Scholar
  28. 28.
    Bard AJ, Faulkner LR, Leddy J, Zoski CG (1980) Electrochemical methods: fundamentals and applications, vol 2. wiley, New YorkGoogle Scholar
  29. 29.
    Boonyasit Y, Chailapakul O, Laiwattanapaisal W (2018) A folding affinity paper-based electrochemical impedance device for cardiovascular risk assessment. doi: Scholar
  30. 30.
    Christopeit T, Gossas T, Danielson UH (2009) Characterization of Ca2+ and phosphocholine interactions with C-reactive protein using a surface plasmon resonance biosensor. Anal Biochem 391(1):39–44. CrossRefPubMedGoogle Scholar
  31. 31.
    Kim E, Kim H-C, Lee SG, Lee SJ, Go T-J, Baek CS, Jeong SW (2011) C-reactive protein-directed immobilization of phosphocholine ligands on a solid surface. Chem Commun 47(43):11900–11902. CrossRefGoogle Scholar
  32. 32.
    Mi LZ, Wang HW, Sui SF (1997) Interaction of rabbit C-reactive protein with phospholipid monolayers studied by microfluorescence film balance with an externally applied electric field. Biophys J 73(1):446–451. CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Suresh MV, Singh SK, Agrawal A (2004) Interaction of calcium-bound C-reactive protein with fibronectin is controlled by pH: IN VIVO IMPLICATIONS. J Biol Chem 279(50):52552–52557. CrossRefPubMedGoogle Scholar
  34. 34.
    Volanakis JE (2001) Human C-reactive protein: expression, structure, and function. Mol Immunol 38(2):189–197. CrossRefPubMedGoogle Scholar
  35. 35.
    Kong B, Kim Y, Choi I (2008) pH-dependent stability of self-assembled monolayers on gold. Bull Kor Chem Soc 29(9):1843–1846CrossRefGoogle Scholar
  36. 36.
    Hutt DA, Leggett GJ (1997) Functionalization of hydroxyl and carboxylic acid terminated self-assembled monolayers. Langmuir 13(10):2740–2748. CrossRefGoogle Scholar
  37. 37.
    Boonkaew S, Chaiyo S, Jampasa S, Rengpipat S, Siangproh W, Chailapakul O (2019) An origami paper-based electrochemical immunoassay for the C-reactive protein using a screen-printed carbon electrode modified with graphene and gold nanoparticles, vol 186. doi:
  38. 38.
    Jampasa S, Siangproh W, Laocharoensuk R, Vilaivan T, Chailapakul O (2018) Electrochemical detection of c-reactive protein based on anthraquinone-labeled antibody using a screen-printed graphene electrode. Talanta 183:311–319. CrossRefPubMedGoogle Scholar
  39. 39.
    Bryan T, Luo X, Bueno PR, Davis JJ (2013) An optimised electrochemical biosensor for the label-free detection of C-reactive protein in blood. Biosens Bioelectron 39(1):94–98. CrossRefPubMedGoogle Scholar
  40. 40.
    Thangamuthu M, Santschi C, J. F. Martin O (2018) Label-free electrochemical immunoassay for C-reactive protein. Biosensors 8 (2):34Google Scholar

Copyright information

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

Authors and Affiliations

  • Chanika Pinyorospathum
    • 1
    • 2
  • Sudkate Chaiyo
    • 2
    • 3
  • Pornpen Sae-ung
    • 1
  • Voravee P. Hoven
    • 1
    • 4
  • Panittha Damsongsang
    • 1
  • Weena Siangproh
    • 5
  • Orawon Chailapakul
    • 1
    • 2
    • 4
    Email author
  1. 1.Department of Chemistry, Faculty of ScienceChulalongkorn UniversityBangkokThailand
  2. 2.Electrochemistry and Optical Spectroscopy Center of Excellence (EOSCE), Department of Chemistry, Faculty of ScienceChulalongkorn UniversityBangkokThailand
  3. 3.The Institute of Biotechnology and Genetic EngineeringChulalongkorn UniversityBangkokThailand
  4. 4.Nanotec-CU Center of Excellence on Food and Agriculture, Department of Chemistry, Faculty of ScienceChulalongkorn UniversityBangkokThailand
  5. 5.Department of Chemistry, Faculty of ScienceSrinakharinwirot UniversityBangkokThailand

Personalised recommendations