Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Preparation of carbon nanotube bioconjugates for biomedical applications

Abstract

Biomedical applications of carbon nanotubes have attracted much attention in recent years. Here, we summarize our previously developed protocols for functionalization and bioconjugation of single-walled carbon nanotubes (SWNTs) for various biomedical applications including biological imaging; using nanotubes as Raman, photoluminescence and photoacoustic labels; sensing using nanotubes as Raman tags and drug delivery. Sonication of SWNTs in solutions of phospholipid-polyethylene glycol (PL-PEG) is our most commonly used protocol of SWNT functionalization. Compared with other frequently used covalent strategies, our non-covalent functionalization protocol largely retains the intrinsic optical properties of SWNTs, which are useful in various biological imaging and sensing applications. Functionalized SWNTs are conjugated with targeting ligands, including peptides and antibodies for specific cell labeling in vitro or tumor targeting in vivo. Radio labels are introduced for tracking and imaging of SWNTs in real time in vivo. Moreover, SWNTs can be conjugated with small interfering RNA (siRNA) or loaded with chemotherapy drugs for drug delivery. These procedures take various times ranging from 1 to 5 d.

Your institute does not have access to this article

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Overview of the protocol.
Figure 2: A scheme showing conjugation of targeting ligands to SWNTs.
Figure 3: A scheme showing radiolabeling of targeting SWNT bioconjugates.
Figure 4: A scheme showing siRNA conjugation to SWNTs through a disulfide bond.
Figure 5: Functionalization of SWNTs by PL-PEG.
Figure 6: Targeting SWNT bioconjugates for cell labeling and Raman imaging.
Figure 7: Radiolabeled SWNTs for in vivo PET imaging and tumor targeting radiolabeled nanotubes.
Figure 8: CXCR4 expression levels on CEM.NKR cells after various treatments.
Figure 9: Doxorubicin (DOX) on functionalized SWNTs for drug delivery.

References

  1. Liu, Z., Tabakman, S., Welsher, K. & Dai, H. Carbon nanotubes in biology and medicine: in vitro and in vivo detection, imaging and drug delivery. Nano Res. 2, 85–120 (2009).

    CAS  Article  Google Scholar 

  2. Bianco, A., Kostarelos, K., Partidos, C.D. & Prato, M. Biomedical applications of functionalised carbon nanotubes. Chem. Commun. 571–577 (2005).

  3. Kam, N.W.S., Jessop, T.C., Wender, P.A. & Dai, H.J. Nanotube molecular transporters: internalization of carbon nanotube-protein conjugates into mammalian cells. J. Am. Chem. Soc. 126, 6850–6851 (2004).

    CAS  Article  Google Scholar 

  4. Liu, Z., Winters, M., Holodniy, M. & Dai, H.J. siRNA delivery into human T cells and primary cells with carbon-nanotube transporters. Angew. Chem. Int. Ed. Engl. 46, 2023–2027 (2007).

    CAS  Article  Google Scholar 

  5. Kam, N.W.S., Liu, Z.A. & Dai, H.J. Carbon nanotubes as intracellular transporters for proteins and DNA: an investigation of the uptake mechanism and pathway. Angew. Chem. Int. Ed. Engl. 45, 577–581 (2006).

    CAS  Article  Google Scholar 

  6. Kam, N.W.S., Liu, Z. & Dai, H. Functionalization of carbon nanotubes via cleavable disulfide bonds for efficient intracellular delivery of siRNA and potent gene silencing. J. Am. Chem. Soc. 127, 12492–12493 (2005).

    CAS  Article  Google Scholar 

  7. Kam, N.W.S. & Dai, H. Carbon nanotubes as intracellular protein transporters: generality and biological functionality. J. Am. Chem. Soc. 127, 6021–6026 (2005).

    CAS  Article  Google Scholar 

  8. Sayes, C.M. et al. Functionalization density dependence of single-walled carbon nanotubes cytotoxicity in vitro . Toxicol. Lett. 161, 135–142 (2006).

    CAS  Article  Google Scholar 

  9. Liu, Z. et al. Circulation and long-term fate of functionalized, biocompatible single-walled carbon nanotubes in mice probed by Raman spectroscopy. Proc. Natl. Acad. Sci. USA 105, 1410–1415 (2008).

    CAS  Article  Google Scholar 

  10. Schipper, M.L. et al. A pilot toxicology study of single-walled carbon nanotubes in a small sample of mice. Nat. Nanotech. 3, 216–221 (2008).

    CAS  Article  Google Scholar 

  11. Liu, Z. et al. Drug delivery with carbon nanotubes for in vivo cancer treatment. Cancer Res. 68, 6652–6660 (2008).

    CAS  Article  Google Scholar 

  12. Heller, D.A., Baik, S., Eurell, T.E. & Strano, M.S. Single-walled carbon nanotube spectroscopy in live cells: towards long-term labels and optical sensors. Adv. Mater. 17, 2793–2799 (2005).

    CAS  Article  Google Scholar 

  13. Leeuw, T.K. et al. Single-walled carbon nanotubes in the intact organism: near-IR imaging and biocompatibility studies in Drosophila. Nano Lett. 7, 2650–2654 (2007).

    CAS  Article  Google Scholar 

  14. Chen, Z. et al. Protein microarrays with carbon nanotubes as multicolor Raman labels. Nat. Biotechnol. 26, 1285–1292 (2008).

    CAS  Article  Google Scholar 

  15. Liu, Z. et al. Multiplexed multi-color Raman imaging of live cells with isotopically modified single walled carbon nanotubes. J. Am. Chem. Soc. 130, 13540–13541 (2008).

    CAS  Article  Google Scholar 

  16. Welsher, K., Liu, Z., Daranciang, D. & Dai, H. Selective probing and imaging of cells with single walled carbon nanotubes as near-infrared fluorescent molecules. Nano Lett. 8, 586–590 (2008).

    CAS  Article  Google Scholar 

  17. de la Zerda, L . et al. Photoacoustic molecular imaging in living mice utilizing targeted carbon nanotubes. Nat. Nanotechnol. 3, 557–562 (2008).

    CAS  Article  Google Scholar 

  18. Zavaleta, C. et al. Noninvasive Raman spectroscopy in living mice for evaluation of tumor targeting with carbon nanotubes. Nano Lett. 8, 2800–2805 (2008).

    CAS  Article  Google Scholar 

  19. Qian, X.M. et al. In vivo tumor targeting and spectroscopic detection with surface-enhanced Raman nanoparticle tags. Nat. Biotechnol. 26, 83–90 (2008).

    CAS  Article  Google Scholar 

  20. Alivisatos, A.P., Gu, W.W. & Larabell, C. Quantum dots as cellular probes. Ann. Rev. Biomed. Eng. 7, 55–76 (2005).

    CAS  Article  Google Scholar 

  21. Liu, Z., Sun, X., Nakayama, N. & Dai, H. Supramolecular chemistry on water-soluble carbon nanotubes for drug loading and delivery. ACS Nano 1, 50–56 (2007).

    Article  Google Scholar 

  22. Kam, N.W.S., O'Connell, M., Wisdom, J.A. & Dai, H. Carbon nanotubes as multifunctional biological transporters and near-infrared agents for selective cancer cell destruction. Proc. Natl. Acad. Sci. USA 102, 11600–11605 (2005).

    CAS  Article  Google Scholar 

  23. Chakravarty, P. et al. Thermal ablation of tumor cells with anti body-functionalized single-walled carbon nanotubes. Proc. Natl. Acad. Sci. USA 105, 8697–8702 (2008).

    CAS  Article  Google Scholar 

  24. Zhang, Z.H. et al. Delivery of telomerase reverse transcriptase small interfering RNA in complex with positively charged single-walled carbon nanotubes suppresses tumor growth. Clin. Cancer Res. 12, 4933–4939 (2006).

    CAS  Article  Google Scholar 

  25. Liu, Z. et al. In vivo biodistribution and highly efficient tumour targeting of carbon nanotubes in mice. Nat. Nanotechnol. 2, 47–52 (2007).

    CAS  Article  Google Scholar 

  26. Niyogi, S. et al. Chemistry of single-walled carbon nanotubes. Acc. Chem. Res. 35, 1105–1113 (2002).

    CAS  Article  Google Scholar 

  27. Rosca, I.D., Watari, F., Uo, M. & Akaska, T. Oxidation of multiwalled carbon nanotubes by nitric acid. Carbon 43, 3124–3131 (2005).

    CAS  Article  Google Scholar 

  28. Tagmatarchis, N. & Prato, M. Functionalization of carbon nanotubes via 1,3-dipolar cycloadditions. J. Mater. Chem. 14, 437–439 (2004).

    CAS  Article  Google Scholar 

  29. Liu, Y. et al. Polyethylenimine-grafted multiwalled carbon nanotubes for secure noncovalent immobilization and efficient delivery of DNA. Angew. Chem. Int. Ed. Engl. 44, 4782–4785 (2005).

    CAS  Article  Google Scholar 

  30. Ross, J.S. & Fletcher, J.A. The HER-2/neu oncogene in breast cancer: prognostic factor, predictive factor, and target for therapy. Stem Cells 16, 413–428 (1998).

    CAS  Article  Google Scholar 

  31. Jin, H. & Varner, J. Integrins: roles in cancer development and as treatment targets. Br. J. Cancer 90, 561–565 (2004).

    CAS  Article  Google Scholar 

  32. Cai, W.B. & Chen, X.Y. Preparation of peptide-conjugated quantum dots for tumor vasculature-targeted imaging. Nat. Protoc. 3, 89–96 (2008).

    CAS  Article  Google Scholar 

  33. Anderson, J., Banerjea, A., Planelles, V. & Akkina, R. Potent suppression of HIV type 1 infection by a short hairpin anti-CXCR4 siRNA. AIDS Res. Hum. Retroviruses 19, 699–706 (2003).

    CAS  Article  Google Scholar 

  34. Novina, C.D. et al. siRNA-directed inhibition of HIV-1 infection. Nat. Med. 8, 681–686 (2002).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

The multiple projects involved here were supported by a Stanford Graduate Fellowship, a Stanford Bio-X grant, CCNE-TR at Stanford University, NIH-NCI R01 CA135109-02 and Ensysce Biosciences Inc. Drs Nadine Wong Shi Kam, Sarunya Bangsaruntip, Xiaowu Tang, Xiaoming Sun, Xiaoyuan Chen, Weibo Cai and Ms Nozomi Nakayama have also contributed in the development of this protocol.

Author information

Authors and Affiliations

Authors

Contributions

Z.L. and H.D. designed and wrote this paper. S.M.T. and Z.C. revised the paper.

Corresponding authors

Correspondence to Zhuo Chen or Hongjie Dai.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Liu, Z., Tabakman, S., Chen, Z. et al. Preparation of carbon nanotube bioconjugates for biomedical applications. Nat Protoc 4, 1372–1381 (2009). https://doi.org/10.1038/nprot.2009.146

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nprot.2009.146

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing