Nano Research

, Volume 11, Issue 8, pp 4005–4016 | Cite as

Surface-enhanced Raman scattering nanosensors for in vivo detection of nucleic acid targets in a large animal model

  • Hsin-Neng Wang
  • Janna K. Register
  • Andrew M. Fales
  • Naveen Gandra
  • Eugenia H. Cho
  • Alina Boico
  • Gregory M. Palmer
  • Bruce Klitzman
  • Tuan Vo-Dinh
Research Article


Although nanotechnology has led to important advances in in vitro diagnostics, the development of nanosensors for in vivo detection remains very challenging. Here, we demonstrated the proof-of-principle of in vivo detection of nucleic acid targets using a promising type of surface-enhanced Raman scattering (SERS) nanosensor implanted in the skin of a large animal model (pig). The in vivo nanosensor used in this study involves the “inverse molecular sentinel” detection scheme using plasmonics-active nanostars, which have tunable absorption bands in the near infrared region of the “tissue optical window”, rendering them efficient as an optical sensing platform for in vivo optical detection. Ex vivo measurements were also performed using human skin grafts to demonstrate the detection of SERS nanosensors through tissue. In this study, a new core–shell nanorattle probe with Raman reporters trapped between the core and shell was utilized as an internal standard system for self-calibration. These results illustrate the usefulness and translational potential of the SERS nanosensor for in vivo biosensing.


nanosensor nanoprobes plasmonics nanostar surface-enhanced Raman scattering (SERS) in vivo sensing 


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This work was sponsored by the Defense Advanced Research Projects Agency (No. HR0011-13-2-0003). The content of the information does not necessarily reflect the position or the policy of the Government, and no official endorsement should be inferred.


  1. [1]
    Campion A.; Kambhampati, P. Surface-enhanced Raman scattering. Chem. Soc. Rev. 1998, 27, 241–250.CrossRefGoogle Scholar
  2. [2]
    Schlücker, S. Surface-enhanced Raman spectroscopy: Concepts and chemical applications. Angew. Chem., Int. Ed. 2014, 53, 4756–4795.CrossRefGoogle Scholar
  3. [3]
    Vo-Dinh, T.; Hiromoto, M. Y. K.; Begun, G. M.; Moody, R. L. Surface-enhanced Raman spectrometry for trace organic analysis. Anal. Chem. 1984, 56, 1667–1670.CrossRefGoogle Scholar
  4. [4]
    Vo-Dinh, T. Surface-enhanced Raman spectroscopy using metallic nanostructures. TrAC-Trend. Anal. Chem. 1998, 17, 557–582.CrossRefGoogle Scholar
  5. [5]
    Vo-Dinh, T.; Fales, A. M.; Griffin, G. D.; Khoury, C. G.; Liu, Y.; Ngo, H.; Norton, S. J.; Register, J. K.; Wang, H. N.; Yuan, H. Plasmonic nanoprobes: From chemical sensing to medical diagnostics and therapy. Nanoscale 2013, 5, 10127–10140.CrossRefGoogle Scholar
  6. [6]
    Ng, V. W. K.; Berti, R.; Lesage, F.; Kakkar, A. Gold: A versatile tool for in vivo imaging. J. Mater. Chem. B 2013, 1, 9–25.CrossRefGoogle Scholar
  7. [7]
    Yuan, H.; Khoury, C. G.; Hwang, H.; Wilson, C. M.; Grant, G. A.; Vo-Dinh, T. Gold nanostars: Surfactant-free synthesis, 3D modelling, and two-photon photoluminescence imaging. Nanotechnology 2012, 23, 075102.CrossRefGoogle Scholar
  8. [8]
    Yuan, H.; Fales, A. M.; Khoury, C. G.; Liu, J.; Vo-Dinh, T. Spectral characterization and intracellular detection of surfaceenhanced Raman scattering (SERS)-encoded plasmonic gold nanostars. J. Raman Spectrosc. 2013, 44, 234–239.CrossRefGoogle Scholar
  9. [9]
    Tian, F. R.; Conde, J.; Bao, C. C.; Chen, Y. S.; Curtin, J.; Cui, D. X. Gold nanostars for efficient in vitro and in vivo real-time SERS detection and drug delivery via plasmonictunable Raman/FTIR imaging. Biomaterials 2016, 106, 87–97.CrossRefGoogle Scholar
  10. [10]
    Fales, A. M.; Yuan, H.; Vo-Dinh, T. Development of hybrid silver-coated gold nanostars for nonaggregated surfaceenhanced Raman scattering. J. Phys. Chem. C 2014, 118, 3708–3715.CrossRefGoogle Scholar
  11. [11]
    Souza, G. R.; Levin, C. S.; Hajitou, A.; Pasqualini, R.; Arap, W.; Miller, J. H. In vivo detection of gold-imidazole selfassembly complexes: NIR-SERS signal reporters. Anal. Chem. 2006, 78, 6232–6237.CrossRefGoogle Scholar
  12. [12]
    Stuart, D. A.; Yuen, J. M.; Shah, N.; Lyandres, O.; Yonzon, C. R.; Glucksberg, M. R.; Walsh, J. T.; Van Duyne, R. P. In vivo glucose measurement by surface-enhanced Raman spectroscopy. Anal. Chem. 2006, 78, 7211–7215.CrossRefGoogle Scholar
  13. [13]
    Qian, X. M.; Peng, X. H.; Ansari, D. O.; Yin-Goen, Q.; Chen, G. Z.; Shin, D. M.; Yang, L.; Young, A. N.; Wang, M. D.; Nie, S. M. In vivo tumor targeting and spectroscopic detection with surface-enhanced Raman nanoparticle tags. Nat. Biotechnol. 2008, 26, 83–90.CrossRefGoogle Scholar
  14. [14]
    Samanta, A.; Maiti, K. K.; Soh, K. S.; Liao, X. J.; Vendrell, M.; Dinish, U. S.; Yun, S. W.; Bhuvaneswari, R.; Kim, H.; Rautela, S. et al. Ultrasensitive near-infrared Raman reporters for SERS-based in vivo cancer detection. Angew. Chem., Int. Ed. 2011, 50, 6089–6092.CrossRefGoogle Scholar
  15. [15]
    Maiti, K. K.; Dinish, U. S.; Fu, C. Y.; Lee, J. J.; Soh, K. S.; Yun, S. W.; Bhuvaneswari, R.; Olivo, M.; Chang, Y. T. Development of biocompatible SERS nanotag with increased stability by chemisorption of reporter molecule for in vivo cancer detection. Biosens. Bioelectron. 2010, 26, 398–403.CrossRefGoogle Scholar
  16. [16]
    McQueenie, R.; Stevenson, R.; Benson, R.; MacRitchie, N.; McInnes, I.; Maffia, P.; Faulds, K.; Graham, D.; Brewer, J.; Garside, P. Detection of inflammation in vivo by surfaceenhanced Raman scattering provides higher sensitivity than conventional fluorescence imaging. Anal. Chem. 2012, 84, 5968–5975.CrossRefGoogle Scholar
  17. [17]
    Dinish, U. S.; Balasundaram, G.; Chang, Y. T.; Olivo, M. Actively targeted in vivo multiplex detection of intrinsic cancer biomarkers using biocompatible SERS nanotags. Sci. Rep. 2014, 4, 4075.CrossRefGoogle Scholar
  18. [18]
    Zavaleta, C. L.; Smith, B. R.; Walton, I.; Doering, W.; Davis, G.; Shojaei, B.; Natan, M. J.; Gambhir, S. S. Multiplexed imaging of surface enhanced Raman scattering nanotags in living mice using noninvasive Raman spectroscopy. Proc. Natl. Acad. Sci. USA 2009, 106, 13511–13516.CrossRefGoogle Scholar
  19. [19]
    Yigit, M. V.; Zhu, L. Y.; Ifediba, M. A.; Zhang, Y.; Carr, K.; Moore, A.; Medarova, Z. Noninvasive MRI-SERS imaging in living mice using an innately bimodal nanomaterial. ACS Nano 2011, 5, 1056–1066.CrossRefGoogle Scholar
  20. [20]
    Kang, H.; Jeong, S.; Park, Y.; Yim, J.; Jun, B. H.; Kyeong, S.; Yang, J. K.; Kim, G.; Hong, S.; Lee, L. P. et al. Near-infrared SERS nanoprobes with plasmonic Au/Ag hollow-shell assemblies for in vivo multiplex detection. Adv. Funct. Mater. 2013, 23, 3719–3727.CrossRefGoogle Scholar
  21. [21]
    Wang, Y. L.; Seebald, J. L.; Szeto, D. P.; Irudayaraj, J. Biocompatibility and biodistribution of surface-enhanced Raman scattering nanoprobes in zebrafish embryos: In vivo and multiplex imaging. ACS Nano 2010, 4, 4039–4053.CrossRefGoogle Scholar
  22. [22]
    Fales, A. M.; Yuan, H.; Vo-Dinh, T. Silica-coated gold nanostars for combined surface-enhanced Raman scattering (SERS) detection and singlet-oxygen generation: A potential nanoplatform for theranostics. Langmuir 2011, 27, 12186–12190.CrossRefGoogle Scholar
  23. [23]
    Conde, J.; Bao, C. C.; Cui, D. X.; Baptista, P. V.; Tian, F. R. Antibody–drug gold nanoantennas with Raman spectroscopic fingerprints for in vivo tumour theranostics. J. Control. Release 2014, 183, 87–93.CrossRefGoogle Scholar
  24. [24]
    Zhang, Y.; Qian, J.; Wang, D.; Wang, Y. L.; He, S. L. Multifunctional gold nanorods with ultrahigh stability and tunability for in vivo fluorescence imaging, SERS detection, and photodynamic therapy. Angew. Chem., Int. Ed. 2013, 52, 1148–1151.CrossRefGoogle Scholar
  25. [25]
    Liu, Y.; Ashton, J. R.; Moding, E. J.; Yuan, H.; Register, J. K.; Fales, A. M.; Choi, J.; Whitley, M. J.; Zhao, X. G.; Qi, Y. et al. A plasmonic gold nanostar theranostic probe for in vivo tumor imaging and photothermal therapy. Theranostics 2015, 5, 946–960.CrossRefGoogle Scholar
  26. [26]
    Schwarzenbach, H.; Hoon, D. S. B.; Pantel, K. Cell-free nucleic acids as biomarkers in cancer patients. Nat. Rev. Cancer 2011, 11, 426–437.CrossRefGoogle Scholar
  27. [27]
    Anker, P.; Lyautey, J.; Lederrey, C.; Stroun, M. Circulating nucleic acids in plasma or serum. Clin. Chim. Acta 2001, 313, 143–146.CrossRefGoogle Scholar
  28. [28]
    Gormally, E.; Caboux, E.; Vineis, P; Hainaut, P. Circulating free DNA in plasma or serum as biomarker of carcinogenesis: Practical aspects and biological significance. Mutat. Res. Rev. Mutat. Res. 2007, 635, 105–117.CrossRefGoogle Scholar
  29. [29]
    Kopreski, M. S.; Benko, F. A.; Kwak, L. W.; Gocke, C. D. Detection of tumor messenger RNA in the serum of patients with malignant melanoma. Clin. Cancer Res. 1999, 5, 1961–1965.Google Scholar
  30. [30]
    Mitchell, P. S.; Parkin, R. K.; Kroh, E. M.; Fritz, B. R.; Wyman, S. K.; Pogosova-Agadjanyan, E. L.; Peterson, A.; Noteboom, J.; O’Briant, K. C.; Allen, A. et al. Circulating microRNAs as stable blood-based markers for cancer detection. Proc. Natl. Acad. Sci. USA 2008, 105, 10513–10518.CrossRefGoogle Scholar
  31. [31]
    Chen, X.; Ba, Y.; Ma, L. J.; Cai, X.; Yin, Y.; Wang, K. H.; Guo, J. G.; Zhang, Y. J.; Chen, J. N.; Guo, X. et al. Characterization of microRNAs in serum: A novel class of biomarkers for diagnosis of cancer and other diseases. Cell Res. 2008, 18, 997–1006.CrossRefGoogle Scholar
  32. [32]
    Wang, H. N.; Fales, A. M.; Vo-Dinh, T. Plasmonics-based SERS nanobiosensor for homogeneous nucleic acid detection. Nanomed. Nanotech. Biol. Med. 2015, 11, 811–814.CrossRefGoogle Scholar
  33. [33]
    Ngo, H. T.; Wang, H. N.; Fales, A. M.; Nicholson, B. P.; Woods, C. W.; Vo-Dinh, T. DNA bioassay-on-chip using SERS detection for dengue diagnosis. Analyst 2014, 139, 5655–5659.CrossRefGoogle Scholar
  34. [34]
    Wang, H. N.; Crawford, B. M.; Fales, A. M.; Bowie, M. L.; Seewaldt, V. L.; Vo-Dinh, T. Multiplexed detection of microRNA biomarkers using SERS-based inverse molecular sentinel (iMS) nanoprobes. J. Phys. Chem. C 2016, 120, 21047–21055.CrossRefGoogle Scholar
  35. [35]
    Wabuyele, M. B.; Vo-Dinh, T. Detection of human immunodeficiency virus type 1DNA sequence using plasmonics nanoprobes. Anal. Chem. 2005, 77, 7810–7815.CrossRefGoogle Scholar
  36. [36]
    Wang, H. N.; Vo-Dinh, T. Multiplex detection of breast cancer biomarkers using plasmonic molecular sentinel nanoprobes. Nanotechnology 2009, 20, 065101.CrossRefGoogle Scholar
  37. [37]
    Gandra, N.; Hendargo, H. C.; Norton, S. J.; Fales, A. M.; Palmer, G. M.; Vo-Dinh, T. Tunable and amplified Raman gold nanoprobes for effective tracking (TARGET): In vivo sensing and imaging. Nanoscale 2016, 8, 8486–8494.CrossRefGoogle Scholar
  38. [38]
    Tian, L. M.; Gandra, N.; Singamaneni, S. Monitoring controlled release of payload from gold nanocages using surface enhanced Raman scattering. ACS Nano 2013, 7, 4252–4260.CrossRefGoogle Scholar
  39. [39]
    Choi, S. W.; Zhang, Y.; Xia, Y. N. A temperature-sensitive drug release system based on phase-change materials. Angew. Chem., Int. Ed. 2010, 49, 7904–7908.CrossRefGoogle Scholar
  40. [40]
    Moon, G. D.; Choi, S. W.; Cai, X.; Li, W. Y.; Cho, E. C.; Jeong, U.; Wang, L. V.; Xia, Y. N. A new theranostic system based on gold nanocages and phase-change materials with unique features for photoacoustic imaging and controlled release. J. Am. Chem. Soc. 2011, 133, 4762–4765.CrossRefGoogle Scholar
  41. [41]
    Register, J. K.; Fales, A. M.; Wang, H. N.; Norton, S. J.; Cho, E. H.; Boico, A.; Pradhan, S.; Kim, J.; Schroeder, T.; Wisniewski, N. A. et al. In vivo detection of SERS-encoded plasmonic nanostars in human skin grafts and live animal models. Anal. Bioanal. Chem. 2015, 407, 8215–8224.CrossRefGoogle Scholar
  42. [42]
    Martinez, K.; Estevez, M. C.; Wu, Y. R.; Phillips, J. A.; Medley, C. D.; Tan, W. H. Locked nucleic acid based beacons for surface interaction studies and biosensor development. Anal. Chem. 2009, 81, 3448–3454.CrossRefGoogle Scholar
  43. [43]
    Ansari, A.; Kuznetsov, S. V. Is hairpin formation in singlestranded polynucleotide diffusion-controlled? J. Phys. Chem. B 2005, 109, 12982–12989.CrossRefGoogle Scholar

Copyright information

© Tsinghua University Press and Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Hsin-Neng Wang
    • 1
    • 2
  • Janna K. Register
    • 1
    • 2
  • Andrew M. Fales
    • 1
    • 2
  • Naveen Gandra
    • 1
    • 2
  • Eugenia H. Cho
    • 3
  • Alina Boico
    • 3
  • Gregory M. Palmer
    • 3
  • Bruce Klitzman
    • 3
  • Tuan Vo-Dinh
    • 1
    • 2
    • 4
  1. 1.Departments of Biomedical EngineeringDuke UniversityDurhamUSA
  2. 2.Fitzpatrick Institute for PhotonicsDuke UniversityDurhamUSA
  3. 3.Medical CenterDuke UniversityDurhamUSA
  4. 4.Departments of ChemistryDuke UniversityDurhamUSA

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