Nano Research

, Volume 9, Issue 6, pp 1701–1708 | Cite as

A simple and novel method for the quantitative detection of 5-hydroxymethylcytosine using carbon nanotube field-effect transistors

  • Fang Yuan
  • Yanyan Deng
  • Wenyu Zhou
  • Min Zhang
  • Zigang Li
Research Article


5-hydroxymethylcytosine (5-hmC) is an important epigenetic derivative of cytosine and quantitative detection of 5-hmC could be used as a reliable biomarker for a variety of human diseases. Current technologies used in 5-hmC detection are complicated and time/cost inefficient. In this work, we report the first application of antibody-functionalized carbon nanotube field-effect transistors (CNT-FETs) in quantitative detection of 5-hmC from mouse tissues. This method achieves facile and ultra-sensitive 5-hmC detection based on electrical performance device and avoids complicated processing for DNA samples. The 5-hmC content percentages of normal mouse cerebrum, cerebellum, spleen, lung, liver, and heart samples presented in the genomic DNA were measured as 0.653, 0.573, 0.002, 0.020, 0.076, and 0.009, respectively, which is consistent with previous reports. This technology could be developed into facile routine 5-hmC monitoring devices for clinic human disease diagnoses.


5-hydroxymethylcytosine carbon nanotube field-effect transistors biosensor 


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  1. [1]
    Wyatt, G. R.; Cohen, S. S. A new pyrimidine base from bacteriophage nucleic acids. Nature 1952, 170, 072–1073.CrossRefGoogle Scholar
  2. [2]
    Penn, N. W.; Suwalski, R.; O’Riley, C.; Bojanowski, K.; Yura, R. The presence of 5-hydroxymethylcytosine in animal deoxyribonucleic acid. Biochem. J. 1972, 126, 81–790.CrossRefGoogle Scholar
  3. [3]
    Kriaucionis, S.; Heintz, N. The nuclear DNA base 5-hydroxymethylcytosine is present in Purkinje neurons and the brain. Science 2009, 324, 29–930.CrossRefGoogle Scholar
  4. [4]
    Tahiliani, M.; Koh, K. P.; Shen, Y. H.; Pastor, W. A.; Bandukwala, H.; Brudno, Y.; Agarwal, S.; Iyer, L. M.; Liu, D. R.; Aravind, L. et al. Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1. Science 2009, 324, 30–935.CrossRefGoogle Scholar
  5. [5]
    Li, W. W.; Liu, M. Distribution of 5-hydroxymethylcytosine in different human tissues. J. Nucleic Acids. 2011, 2011, 870726.CrossRefGoogle Scholar
  6. [6]
    Feinberg, A. P.; Ohlsson, R; Henikoff, S. The epigenetic progenitor origin of human cancer. Nat. Rev. Genet. 2006, 7, 1–33.CrossRefGoogle Scholar
  7. [7]
    Suzuki, M. M.; Bird, A. DNA methylation landscapes: Provocative insights from epigenomics. Nat. Rev. Genet. 2008, 9, 65–476.Google Scholar
  8. [8]
    Kroeze, L. I.; van der Reijden, B. A.; Jansen, J. H. 5-Hydroxymethylcytosine: An epigenetic mark frequently deregulated in cancer. Biochim. Biophys. Acta 2015, 1855, 44–154.Google Scholar
  9. [9]
    Szulwach, K. E.; Li, X. K.; Li, Y. J.; Song, C. X.; Wu, H.; Dai, Q.; Irier, H.; Upadhyay, A. K.; Gearing, M.; Levey, A. I. et al. 5-hmC-mediated epigenetic dynamics during postnatal neurodevelopment and aging. Nat. Neurosci. 2011, 14, 607–1616.CrossRefGoogle Scholar
  10. [10]
    Sherwani, S. I.; Khan, H. A. Role of 5-hydroxymethylcytosine in neurodegeneration. Gene 2015, 570, 7–24.CrossRefGoogle Scholar
  11. [11]
    Al-Mahdawi, S.; Virmouni, S. A.; Pook, M. A. The emerging role of 5-hydroxymethylcytosine in neurodegenerative diseases. Front. Neurosci. 2014, 8, 397.CrossRefGoogle Scholar
  12. [12]
    Clark, S. J.; Harrison, J.; Paul, C. L.; Frommer, M. High sensitivity mapping of methylated cytosines. Nucleic Acids Res. 1994, 22, 990–2997.Google Scholar
  13. [13]
    Cokus, S. J.; Feng, S. H.; Zhang, X. Y.; Chen, Z. G.; Merriman, B.; Haudenschild, C. D.; Pradhan, S.; Nelson, S. F.; Pellegrini, M.; Jacobsen, S. E. Shotgun bisulphite sequencing of the Arabidopsis genome reveals DNA methylation patterning. Nature 2008, 452, 15–219.CrossRefGoogle Scholar
  14. [14]
    Le, T.; Kim, K. P; Fan, G. P.; Faull, K. F. A sensitive mass spectrometry method for simultaneous quantification of DNA methylation and hydroxymethylation levels in biological samples. Anal. Biochem. 2011, 412, 03–209.CrossRefGoogle Scholar
  15. [15]
    Globisch, D.; Münzel, M.; Müller, M.; Michalakis, S.; Wagner, M.; Koch, S.; Brückl, T.; Biel, M.; Carell, T. Tissue distribution of 5-hydroxymethylcytosine and search for active demethylation intermediates. PLoS One 2010, 5–e15367.Google Scholar
  16. [16]
    Ito, S.; Shen, L.; Dai, Q.; Wu, S. C.; Collins, L. B.; Swenberg, J. A.; He, C.; Zhang, Y. Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine. Science 2011, 333, 300–1303.CrossRefGoogle Scholar
  17. [17]
    Song, C. X.; Szulwach, K. E.; Fu, Y.; Dai, Q.; Yi, C. Q.; Li, X. K.; Li, Y. J.; Chen, C. H.; Zhang, W.; Jian, X. et al. Selective chemical labeling reveals the genome-wide distribution of 5-hydroxymethylcytosine. Nat. Biotechnol. 2011, 29, 8–72.CrossRefGoogle Scholar
  18. [18]
    Pastor, W. A.; Pape, U. J.; Huang, Y.; Henderson, H. R.; Lister, R.; Ko, M.; McLoughlin, E. M.; Brudno, Y.; Mahapatra, S.; Kapranov, P. et al. Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells. Nature 2011, 473, 94–397.CrossRefGoogle Scholar
  19. [19]
    Szwagierczak, A.; Bultmann, S.; Schmidt, C. S.; Spada, F.; Leonhardt, H. Sensitive enzymatic quantification of 5-hydroxymethylcytosine in genomic DNA. Nucleic Acids Res. 2010, 38. e181.Google Scholar
  20. [20]
    Terragni, J.; Bitinaite, J.; Zheng, Y.; Pradhan, S. Biochemical characterization of recombinant ß-glucosyltransferase and analysis of global 5-hydroxymethylcytosine in unique genomes. Biochemistry 2012, 51, 009–1019.Google Scholar
  21. [21]
    Kinney, S. M.; Chin, H. G.; Vaisvila, R.; Bitinaite, J.; Zheng, Y.; Estève, P. O.; Feng, S. H.; Stroud, H.; Jacobsen, S. E.; Pradhan, S. Tissue-specific distribution and dynamic changes of 5-hydroxymethylcytosine in mammalian genomes. J. Biol. Chem. 2011, 286, 4685–24693.CrossRefGoogle Scholar
  22. [22]
    Höbartner, C. Enzymatic labeling of 5-hydroxymethylcytosine in DNA. Angew. Chem., Int. Ed. 2011. 50, 4268–4270Google Scholar
  23. [23]
    Harris. P. J. F. Carbon Nanotubes and Related Structures: New Materials for the Twenty-First Century; Cambridge University Press: Cambridge, 2001.Google Scholar
  24. [24]
    Dai, H. J. Carbon nanotubes: Opportunities and challenges. Surf. Sci. 2002, 500, 18–241.CrossRefGoogle Scholar
  25. [25]
    Katz, E.; Willner, I. Biomolecule functionalized carbon nanotubes: Applications in nanobioelectronics. ChemPhysChem 2004, 5, 084–1104.Google Scholar
  26. [26]
    Daniel, S.; Rao, T. P.; Rao, K. S.; Rani, S. U.; Naidu, G. R. K.; Lee, H. Y.; Kawai, T. A review of DNA functionalized/ grafted carbon nanotubes and their characterization. Sens. Act. B 2007, 122, 72–682.CrossRefGoogle Scholar
  27. [27]
    Drouvalakis, K. A.; Bangsaruntip, S.; Wolfgang, H.; Kozar, L. G.; Utz, P. J.; Dai, H. J. Peptide-coated nanotube-based biosensor for the detection of disease-specific autoantibodies in human serum. Biosens. Bioelectron. 2008, 23, 413–1421.Google Scholar
  28. [28]
    Shim, M.; Kam, N. W. S.; Chen, R.; Li, Y. M.; Dai, H. J. Functionalization of carbon nanotubes for biocompatibility and biomolecular recognition. Nano Lett. 2002, 2, 85–288.CrossRefGoogle Scholar
  29. [29]
    Javey, A.; Guo, J.; Wang, Q.; Lundstrom, M.; Dai, H. J. Ballistic carbon nanotube field-effect transistors. Nature 2003, 424, 54–657.CrossRefGoogle Scholar
  30. [30]
    Byon, H. R.; Choi, H. C. Network single-walled carbon nanotube-field effect transistors (SWNT-FETs) with increased Schottky contact area for highly sensitive biosensor applications. J. Am. Chem. Soc. 2006, 128, 188–2189.CrossRefGoogle Scholar
  31. [31]
    Chen, R. J.; Bangsaruntip, S.; Drouvalakis, K. A.; Kam, N. W. S.; Shim, M.; Li, Y. M.; Kim, W.; Utz, P. J.; Dai, H. J. Noncovalent functionalization of carbon nanotubes for highly specific electronic biosensors. Proc. Natl. Acad. Sci. USA 2003, 100, 984–4989.Google Scholar
  32. [32]
    Li, C.; Curreli, M.; Lin, H.; Lei, B.; Ishikawa, F. N.; Datar, R.; Cote, R. J.; Thompson, M. E.; Zhou, C. W. Complementary detection of prostate-specific antigen using In2O3 nanowires and carbon nanotubes. J. Am. Chem. Soc. 2005, 127, 2484–12485.Google Scholar
  33. [33]
    Park, D. W.; Kim, Y. H.; Kim, B. S.; So, H. M.; Won, K.; Lee, J. O; Kong, K. J.; Chang, H. Detection of tumor markers using single-walled carbon nanotube field effect transistors. J. Nanosci. Nanotechnol. 2006, 6, 499–3502.CrossRefGoogle Scholar
  34. [34]
    Takeda, S.; Sbagyo, A.; Sakoda, Y.; Ishii, A.; Sawamura, M.; Sueoka, K.; Kida, H.; Mukasa, K.; Matsumoto, K. Application of carbon nanotubes for detecting anti-hemagglutinins based on antigen-antibody interaction. Biosens. Bioelectron. 2005, 21, 01–205.CrossRefGoogle Scholar
  35. [35]
    Veetil, J. V.; Ye, K. M. Development of immunosensors using carbon nanotubes. Biotechnol. Prog. 2007, 23, 17–531.Google Scholar
  36. [36]
    Liu, Z.; Zhao, J. W.; Xu, W. Y.; Qian, L.; Nie, S. H.; Cui, Z. Effect of surface wettability properties on the electrical properties of printed carbon nanotube thin-film transistors on SiO2/Si substrates. ACS Appl. Mater. Interfaces 2014, 6, 997–10004.Google Scholar
  37. [37]
    Maehashi, K.; Katsura, T.; Kerman, K.; Takamura, Y.; Matsumoto, K.; Tamiya, E. Label-free protein biosensor based on aptamer-modified carbon nanotube field-effect transistors. Anal. Chem. 2007, 79, 82–787.Google Scholar
  38. [38]
    Wang, C.; Zhang, J. L.; Ryu, K.; Badmaev, A.; De Arco, L. G.; Zhou, C. W. Wafer-scale fabrication of separated carbon nanotube thin-film transistors for display applications. Nano Lett. 2009, 9, 285–4291.Google Scholar
  39. [39]
    Tsang, S. C.; Davis, J. J.; Green, M. L. H.; Hill, H. A. O.; Leung, Y. C.; Sadler, P. J. Immobilization of small proteins in carbon nanotubes: High-resolution transmission electron microscopy study and catalytic activity. J. Chem. Soc., Chem. Commun. 1995, 1803–1804.Google Scholar
  40. [40]
    Tsang, S. C.; Guo, Z. J.; Chen, Y. K.; Green, M. L. H.; Hill, H. A. O.; Hambley, T. W.; Sadler, P. J. Immobilization of platinated and iodinated oligonucleotides on carbon nanotubes. Angew. Chem., Int. Ed. 1997, 36, 198–2200.CrossRefGoogle Scholar
  41. [41]
    Guo, Z. J.; Sadler, P. J.; Tsang, S. C. Immobilization and visualization of DNA and proteins on carbon nanotubes. Adv. Mater. 1998, 10, 01–703.Google Scholar
  42. [42]
    Balavoine, F.; Schultz, P.; Richard, C.; Mallouh, V.; Ebbeson, T. W.; Mioskowski, C. Helical crystallization of proteins on carbon nanotubes: A first step towards the development of new biosensors. Angew. Chem., Int. Ed. 1999, 38, 912–1915.CrossRefGoogle Scholar
  43. [43]
    Chen, R. J.; Zhang, Y. G.; Wang, D. W.; Dai, H. J. Noncovalent sidewall functionalization of single-walled carbon nanotubes for protein immobilization. J. Am. Chem. Soc. 2001, 123, 838–3839.Google Scholar
  44. [44]
    Lee, C. S.; Baker, S. E.; Marcus, M. S.; Yang, W. S.; Eriksson, M. A.; Hamers, R. J. Electrically addressable biomolecular functionalization of carbon nanotube and carbon nanofiber electrodes. Nano Lett. 2004, 4, 713–1716.CrossRefGoogle Scholar
  45. [45]
    Hermanson, G. T. Bioconjugate Techniques; Academic Press: San Diego, 1996.Google Scholar
  46. [46]
    Thordarson, P.; Atkin, R.; Kalle, W. H. J; Warr, G. G.; Braet, F. Developments in using scanning probe microscopy to study molecules on surfaces—From thin films and singlemolecule conductivity to drug-living cell interactions. ChemInform 2006, 37. DOI: 10.1002/chin.200639277.Google Scholar
  47. [47]
    Katz, E. Application of bifunctional reagents for immobilization of proteins on a carbon electrode surface: Oriented immobilization of photosynthetic reaction centers. J. Electroanal. Chem. 1994, 365, 57–164.CrossRefGoogle Scholar
  48. [48]
    Jaegfeldt, H.; Kuwana, T.; Johansson, G. Electrochemical stability of catechols with a pyrene side chain strongly adsorbed on graphite electrodes for catalytic oxidation of dihydronicotinamide adenine dinucleotide. J. Am. Chem. Soc. 1983, 105, 805–1814.CrossRefGoogle Scholar

Copyright information

© Tsinghua University Press and Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  1. 1.School of Chemical Biology & BiotechnologyPeking UniversityShenzhenChina
  2. 2.School of Electronic and Computer EngineeringPeking UniversityShenzhenChina
  3. 3.Shenzhen Thin Film Transistor and Advanced Display LabShenzhenChina
  4. 4.Shenzhen Second People’s HospitalShenzhenChina

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