Enzyme-Immobilized CNT Network Probe for In Vivo Neurotransmitter Detection

  • Gi-Ja Lee
  • Seok Keun Choi
  • Samjin Choi
  • Ji Hye Park
  • Hun-Kuk ParkEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 743)


Glutamate is the principal excitatory neurotransmitter in the brain, and its excessive release plays a key role in neuronal death associated with a wide range of neural disorders. Real-time monitoring of extracellular glutamate levels would be very helpful in understanding the excitotoxic process of neurotransmitters on brain injury. Toward the detection of l-glutamate, we describe in this chapter the preparation of carbon nanotube (CNT) network probes with immobilized l-glutamate oxidase (GLOD) by using a non-covalent functionalized method. Such GOLD-CNT network probes are evaluated with real-time electronic responses corresponding to standard glutamate solutions in vitro and a 11-vessel occlusion (11 VO) rat model in vivo. The ultrahigh sensitivity, selectivity, and fast response time of GLOD-CNT network probes are greatly promising for the real-time electronic detection of extracellular glutamate levels in brain.

Key words

Glutamate CNT network probe in vivo real-time detection GLOD immobilization 



This study was supported by the research fund from the Ministry of Commerce, Industry, and Energy (MOCIE Grant #10017190-2007-31), and Bio R&D program through MEST (2010-0019912). We greatly appreciate Mr. B.Y. Lee, Mr. D.H. Son, and Prof. S. Hong at Seoul National University who provided the CNT probes.


  1. 1.
    Gruner, G. (2006) Carbon nanotube transistors for biosensing applications. Anal. Bioanal. Chem. 384, 322–335.CrossRefGoogle Scholar
  2. 2.
    Tans, S. J., Verschueren, A. R. M., and Dekker, C. (1998) Room-temperature transistor based on a single carbon nanotube. Nature (London) 393, 49–52.CrossRefGoogle Scholar
  3. 3.
    Martel, R., Schmidt, T., Shea, H. R., Hertel, T., and Avouris, P. (1998) Single- and multi-wall carbon nanotube field-effect transistors. Appl. Phys. Lett. 73, 2447–2449.CrossRefGoogle Scholar
  4. 4.
    Collins, P. G., Bradley, K., Ishigami, M., and Zetti, A. (2000) Extreme oxygen sensitivity of electronic properties of carbon nanotubes. Science 287, 1801–1804.CrossRefGoogle Scholar
  5. 5.
    Bradley, K., Jhi, S. H., Collins, P. G., Hone, J., Cohen, M. L., Louie, S. G., and Zettl, A. (2002) Is the intrinsic thermoelectric power of carbon nanotubes positive? Phys. Rev. Lett. 85, 4361–4364.CrossRefGoogle Scholar
  6. 6.
    Kong, J., Franklin, N. R., Zhou, C., Chapline, M. G., Peng, S., Cho, K., and Dai, H. (2000) Nanotube molecular wires as chemical sensors. Science 287, 622–625.CrossRefGoogle Scholar
  7. 7.
    Star, A., Gabriel, J. C. P., Bradley, K., and Gruner, G. (2003) Electronic detection of specific protein binding using nanotube FET devices. Nano Lett. 3, 459–463.CrossRefGoogle Scholar
  8. 8.
    Artyukhin, B. A., Stadermann, M., Friddle, R. W., Stroeve, P., Bakajin, O., and Noy, A. (2006) Controlled electrostatic gating of carbon nanotube FET devices. Nano Lett. 6, 2080–2085.CrossRefGoogle Scholar
  9. 9.
    Burghard, M., and Balasubramanian, K. (2005) Chemically functionalized carbon nanotubes. Small 1, 180–192.CrossRefGoogle Scholar
  10. 10.
    Chen, R. J., Zhang, Y., Wang, D., and Dai, H. (2001) Noncovalent sidewall functionalization of single-walled carbon nanotubes for protein immobilization. J. Am. Chem. Soc. 123, 3838–3839.CrossRefGoogle Scholar
  11. 11.
    Shim, M., Kam, N. W. S., Chen, R. J., Li, Y., and Dai, H. (2002) Functionalization of carbon nanotubes for biocompatibility and biomolecular recognition. Nano Lett. 2, 285–288.CrossRefGoogle Scholar
  12. 12.
    Besteman, K., Lee, J. O., Wiertz, F. G. M., Heering, H. A., and Dekker, C. (2003) Enzyme-coated carbon nanotubes as single-molecule biosensors. Nano Lett. 3, 727–730.CrossRefGoogle Scholar
  13. 13.
    Zhao, H., Asai, S., Kohno, T., and Ishikawa, K. (1998) Effects of brain temperature on CBF thresholds for extracellular glutamate release and reuptake in the striatum in a rat model of graded global ischemia. Neuroreport 9, 3183–3188.CrossRefGoogle Scholar
  14. 14.
    Hossmann, K. A. (1994) Viability thresholds and the penumbra of focal ischemia. Ann. Neurol. 36, 557–565.CrossRefGoogle Scholar
  15. 15.
    Busto, R., Globus, M. Y., Dietrich, W. D., Martinez, E., Valdes, I., and Ginsberg, M. D. (1989) Effect of mild hypothermia on ischemia-induced release of neurotransmitters and free fatty acids in rat brain. Stroke 20, 904–910.Google Scholar
  16. 16.
    Benveniste, H., Drejer, J., Schousboe, A., and Diemer, N. H. (1984) Elevation of the extracellular concentrations of glutamate and aspartate in rat hippocampus during transient cerebral ischemia monitored by intracerebral microdialysis. J. Neurochem. 43, 1369–1374.CrossRefGoogle Scholar
  17. 17.
    Wahl, F., Obrenovitch, T. P., Hardy, A. M., Plotkine, M., Boulu, R., and Symon, L. (1994) Extracellular glutamate during focal cerebral ischaemia in rats: Time course and calcium dependency. J. Neurochem. 63, 1003–1011.CrossRefGoogle Scholar
  18. 18.
    Obrenovitch, T. P., Urenjak, J., Richards, D. A., Ueda, Y., Curzon, G., and Symon, L. (1993) Extracellular neuroactive amino acids in the rat striatum during ischaemia: Comparison between penumbral conditions and ischaemia with sustained anoxic depolarization. J. Neurochem. 61, 178–186.CrossRefGoogle Scholar
  19. 19.
    Takagi, K., Ginsberg, M. D., Globus, M. Y., Dietrich, W. D., Martinez, E., and Kraydieh, S. (1993) Changes in amino acid neurotransmitters and cerebral blood flow in the ischemic penumbral region following middle cerebral artery occlusion in the rat: Correlation with histopathology. J. Cerebr. Blood Flow Metab. 13, 575–585.CrossRefGoogle Scholar
  20. 20.
    Hirota, H., Katayama, Y., Kawamata, T., Kano, T., and Tsubokawa, T. (1995) Inhibition of the high-affinity glutamate uptake system facilitates the massive potassium flux during cerebral ischaemia in vivo. Neurol. Res. 17, 94–96.Google Scholar
  21. 21.
    Katayama, Y., Kawamata, T., Tamura, T., Hovda, D. A., Becker, D. P., and Tsubokawa, T. (1991) Calcium-dependent glutamate release concomitant with massive potassium flux during cerebral ischemia in vivo. Brain Res. 558, 136–140.CrossRefGoogle Scholar
  22. 22.
    Albery, W. J., Boutelle, M. G., and Galley, P. T. (1992) The dialysis electrode – A new method for in vivo monitoring. J. Chem. Soc. Chem. Commun. 12, 900–901.CrossRefGoogle Scholar
  23. 23.
    Lee, M., Im, J., Lee, B. Y., Myung, S., Kang, J., Huang, L., Kwon, Y. K., and Hong, S. (2006) Linker-free directed assembly of high-performance integrated devices based on nanotubes and nanowires. Nat. Nanotechnol. 1, 66–71.CrossRefGoogle Scholar
  24. 24.
    Lee, G. J., Lim, J. E., Park, J. H., Choi, S. K., Hong, S., and Park, H. K. (2009) Neurotransmitter detection by enzyme-immobilized CNT-FET. Curr. Appl. Phys. 9, S25–S28.CrossRefGoogle Scholar
  25. 25.
    Choi, S. K., Lee, G. J., Lim, J. E., Eo, Y. H., Kang, S. W., Han, J. H., Kim, B. S., Oh, B. S., and Park, H. K. (2008) Development of animal model for cerebral ischemia. Korean J. Integr. Med. 3, 11–16.Google Scholar
  26. 26.
    Caragine, L. P., Park, H. K., Diaz, F. G., and And Phillis, J. W. (1998) Real-time measurement of ischemia-evoked glutamate release in the cerebral cortex of four and eleven vessel rat occlusion models. Brain Res. 793, 255–264.CrossRefGoogle Scholar
  27. 27.
    Lee, G. J., Choi, S. K., Eo, Y. H., Kang, S. W., Choi, S., Park, J. H., Lim, J. E., Hong, K. W., Jin, H. S., Oh, B. S., and Park, H. K. (2009) The effect of extracellular glutamate release on repetitive transient ischemic injury in global ischemia model. Korean J. Physiol. Pharmacol. 13, 23–26.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Gi-Ja Lee
    • 1
    • 2
  • Seok Keun Choi
    • 3
  • Samjin Choi
    • 1
    • 2
  • Ji Hye Park
    • 1
    • 2
  • Hun-Kuk Park
    • 1
    • 2
    • 4
    Email author
  1. 1.Department of Biomedical EngineeringCollege of Medicine, Kyung Hee UniversitySeoulRepublic of Korea
  2. 2.Healthcare Industry Research Institute, Kyung Hee UniversitySeoulRepublic of Korea
  3. 3.Department of NeurosurgeryKyung Hee University Medical CenterSeoulRepublic of Korea
  4. 4.Program of Medical EngineeringKyung Hee UniversitySeoulRepublic of Korea

Personalised recommendations