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.

  • Article
  • Published:

Decoupling competing surface binding kinetics and reconfiguration of receptor footprint for ultrasensitive stress assays

Nature Nanotechnology volume 10, pages 899–907 (2015)Cite this article

Subjects

Abstract

Cantilever arrays have been used to monitor biochemical interactions and their associated stress. However, it is often necessary to passivate the underside of the cantilever to prevent unwanted ligand adsorption, and this process requires tedious optimization. Here, we show a way to immobilize membrane receptors on nanomechanical cantilevers so that they can function without passivating the underlying surface. Using equilibrium theory, we quantitatively describe the mechanical responses of vancomycin, human immunodeficiency virus type 1 antigens and coagulation factor VIII captured on the cantilever in the presence of competing stresses from the top and bottom cantilever surfaces. We show that the area per receptor molecule on the cantilever surface influences ligand–receptor binding and plays an important role on stress. Our results offer a new way to sense biomolecules and will aid in the creation of ultrasensitive biosensors.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Nanomechanics of a SAM to investigate the complex interplay between the Au (top) and Si (bottom) surfaces of the cantilever and ligand–receptor binding interactions.
Figure 2: Nanomechanical and SPR quantitation of surface binding reactions.
Figure 3: Quantitation of ligand–receptor interactions at fixed VSR concentrations using nanomechanical and SPR assays.
Figure 4: Investigating the effect of surface footprint (area per receptor) on ligand binding.
Figure 5: Quantitative monitoring of protein interactions using mechanical and SPR biosensors.
Figure 6: Nanomechanical quantitation of clotting factors for bleeding-related disorders.

Similar content being viewed by others

References

  1. Son, K., Guasto, J. S. & Stocker, R. Bacteria can exploit a flagellar buckling instability to change direction. Nature Phys. 9, 494–498 (2013).

    Article  CAS  Google Scholar 

  2. Lele, P. P., Hosu, B. G. & Berg, H. C. Dynamics of mechanosensing in the bacterial flagellar motor. Proc. Natl Acad. Sci. USA 110, 11839–11844 (2013).

    Article  CAS  Google Scholar 

  3. Gebhardt, J. C. & Rief, M. Biochemistry force signaling in biology. Science 324, 1278–1280 (2009).

    Article  CAS  Google Scholar 

  4. Arlett, J. L., Myers, E. B. & Roukes, M. L. Comparative advantages of mechanical biosensors. Nature Nanotech. 6, 203–215 (2011).

    Article  CAS  Google Scholar 

  5. Müller, D. J. & Dufrêne, Y. F. Atomic force microscopy as a multifunctional molecular toolbox in nanobiotechnology. Nature Nanotech. 3, 261–269 (2008).

    Article  Google Scholar 

  6. Huber, F., Lang, H. P., Backmann, N., Rimoldi, D. & Gerber, C. Direct detection of a BRAF mutation in total RNA from melanoma cells using cantilever arrays. Nature Nanotech. 8, 125–129 (2013).

    Article  CAS  Google Scholar 

  7. McKendry, R. A. et al. Multiple label-free biodetection and quantitative DNA-binding assays on a nanomechanical cantilever array. Proc. Natl Acad. Sci. USA 99, 9783–9788 (2002).

    Article  CAS  Google Scholar 

  8. Wu, G. H. et al. Bioassay of prostate-specific antigen (PSA) using microcantilevers. Nature Biotechnol. 19, 856–860 (2001).

    Article  CAS  Google Scholar 

  9. Fritz, J. et al. Translating biomolecular recognition into nanomechanics. Science 288, 316–318 (2000).

    Article  CAS  Google Scholar 

  10. Berger, R. et al. Surface stress in the self-assembly of alkanethiols on gold. Science 276, 2021–2024 (1997).

    Article  CAS  Google Scholar 

  11. Gfeller, K. Y., Nugaeva, N. & Hegner, M. Micromechanical oscillators as rapid biosensor for the detection of active growth of Escherichia coli. Biosens. Bioelectron. 21, 528–533 (2005).

    Article  CAS  Google Scholar 

  12. Longo, G. et al. Rapid detection of bacterial resistance to antibiotics using AFM cantilevers as nanomechanical sensors. Nature Nanotech. 8, 522–526 (2013).

    Article  CAS  Google Scholar 

  13. Ndieyira, J. W. et al. Surface-stress sensors for rapid and ultrasensitive detection of active free drugs in human serum. Nature Nanotech. 9, 225–232 (2014).

    Article  CAS  Google Scholar 

  14. Ndieyira, J. W. et al. Nanomechanical detection of antibiotic–mucopeptide binding in a model for superbug drug resistance. Nature Nanotech. 3, 691–696 (2008).

    Article  CAS  Google Scholar 

  15. Dupres, V. et al. Nanoscale mapping and functional analysis of individual adhesins on living bacteria. Nature Methods 2, 515–520 (2001).

    Article  Google Scholar 

  16. Zhang, J. et al. Rapid and label-free nanomechanical detection of biomarker transcripts in human RNA. Nature Nanotech. 1, 214–220 (2006).

    Article  CAS  Google Scholar 

  17. Kosaka, P. M. et al. Detection of cancer biomarkers in serum using a hybrid mechanical and optoplasmonic nanosensor. Nature Nanotech. 9, 1047–1053 (2014).

    Article  CAS  Google Scholar 

  18. Boisen, A., Dohn, S., Keller, S. S., Schmid, S. & Tenje, M. Cantilever-like micromechanical sensors. Rep. Prog. Phys. 74, 036101 (2011).

    Article  Google Scholar 

  19. Burg, T. P. et al. Weighing of biomolecules, single cells and single nanoparticles in fluid. Nature 446, 1066–1069 (2007).

    Article  CAS  Google Scholar 

  20. Shu, W. et al. DNA molecular motor driven micromechanical cantilever arrays. J. Am. Chem. Soc. 127, 17054–17060 (2005).

    Article  CAS  Google Scholar 

  21. Backmann, N. et al. A label-free immunosensor array using single-chain antibody fragments. Proc. Natl Acad. Sci. USA 102, 14587–14592 (2005).

    Article  CAS  Google Scholar 

  22. Mukhopadhyay, R. et al. Cantilever sensor for nanomechanical detection of specific protein conformations. Nano Lett. 5, 2385–2388 (2005).

    Article  CAS  Google Scholar 

  23. Nieto, M. & Perkins, H. R. Modifications of acyl–D-alanyl–D-alanine terminus affecting complex-formation with vancomycin. Biochem. J. 123, 773–787 (1971).

    Article  CAS  Google Scholar 

  24. Williams, D. H., Maguire, A. J., Tsuzuki, W. & Westwell, M. S. An analysis of the origins of a cooperative binding energy of dimerisation. Science 280, 711–714 (1998).

    Article  CAS  Google Scholar 

  25. Sieradzki, K. et al. The development of vancomycin resistance in a patient with methicillin-resistant Staphylococcus aureus infection. N. Engl. J. Med. 340, 517–523 (1999).

    Article  CAS  Google Scholar 

  26. Barna, J. C. & Williams, D. H. The structure and mode of action of glycopeptides antibiotics of the vancomycin group. Annu. Rev. Microbiol. 38, 339–357 (1984).

    Article  CAS  Google Scholar 

  27. West, R. Inorganic chemistry: two-armed silicon. Nature 485, 49–50 (2012).

    Article  CAS  Google Scholar 

  28. Hamers, R. J. Formation and characterization of organic monolayers on semiconductor surfaces. Annu. Rev. Anal. Chem. 1, 707–736 (2008).

    Article  CAS  Google Scholar 

  29. Haas, A. The chemistry of silicon–sulfur compounds. Angew. Chem. Int. Ed. Engl. 4, 1014–1023 (1965).

    Article  CAS  Google Scholar 

  30. Love, J. C. et al. Self-assembled monolayers of thiolates on metals as a form of nanotechnology. Chem. Rev. 289, 1103–1169 (2005).

    Article  Google Scholar 

  31. Kolomenskii, A. A., Gershon, P. D. & Schuessler, H. A. Sensitivity and detection limit of concentration and adsorption measurements by laser-induced surface-plasmon resonance. Appl. Opt. 36, 6539–6547 (1997).

    Article  CAS  Google Scholar 

  32. Stenberg, E., Persson, B., Roos, H. & Urbaniczky, C. Quantitative determination of surface concentration of protein surface plasmon resonance using radiolabeled proteins. J. Colloid Interface Sci. 143, 513–526 (1991).

    Article  CAS  Google Scholar 

  33. Morita, M., Ohmi, T., Hasegawa, E., Kawakami, M. & Ohwada, M. Growth of native oxide on a silicon surface. J. Appl. Phys. 68, 1272–1281 (1990).

    Article  CAS  Google Scholar 

  34. Prime, K. L. & Whitesides, G. M. Self-assembled organic monolayers—model systems for studying adsorption of proteins at surfaces. Science 252, 1164–1167 (1991).

    Article  CAS  Google Scholar 

  35. Prime, K. L. & Whitesides, G. M. Adsorption of proteins onto surfaces containing end-attached oligo(ethylene oxide)—a model system using self-assembled monolayers. J. Am. Chem. Soc. 115, 10714–10721 (1993).

    Article  CAS  Google Scholar 

  36. Hamers-Casterman, C. et al. Naturally occurring antibodies devoid of light chains. Nature 363, 446–448 (1993).

    Article  CAS  Google Scholar 

  37. Hultberg, A. et al. Lactobacillli expressing llama VHH fragments neutralise Lactococcus phages. BMC Biotechnology 84, 58 (2007).

    Article  Google Scholar 

  38. Strokappe, N. et al. Llama antibodies recognizing various epitopes of the CD4bs neutralize a broad range of HIV-1 subtypes A, B, and C. PLoS One 7, e33298 (2012).

    Article  CAS  Google Scholar 

  39. McCoy, L. E. et al. Potent and broad neutralization of HIV-1 by a llama antibody elicited by immunization. J. Exp. Med. 209, 1091–1103 (2012).

    Article  CAS  Google Scholar 

  40. McCoy, L. E. et al. Molecular evolution of broadly neutralizing llama antibodies to the CD4-binding site of HIV-1. PLoS Pathog 10, e1004552 (2014).

    Article  Google Scholar 

  41. McCoy, L. E. et al. Broadly neutralizing VHH against HIV-1. Patent application WO 2013036130 A1 (EP2753644A1, US201501158934).

  42. Zhou, F. et al. Sensitive sandwich ELISA based on a gold nanoparticle layer for cancer detection. Analyst 137, 1779–1784 (2012).

    Article  CAS  Google Scholar 

  43. Liu, J., Bartesaghi, A., Borgnia, M. J., Sapiro, G. & Subramaniam, S. Molecular architecture of native HIV-1 gp120 trimers. Nature 455, 109–113 (2008).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank the EPSRC Grand Challenge in Nanotechnology for Healthcare (EP/G0620064/1), I-sense EPSRC IRC in Early Warning Sensing Systems for Infectious Diseases (EP/G062064/1), Royal Society (RS), Targanta Therapeutics, Bio Nano Consulting (BNC), the European Union FP7 Project VSMMART Nano (managed by BNC) for funding. The authors also thank J. Russat (London Centre for Nanotechnology), S. Sivachelvam (London Centre for Nanotechnology), M. Rehak (Sphere Fluidics, UK), R.A. Weiss (University College London) C.T. Verrips (QVQuality, Utrecht), T. Philips (Utrecht University), M. Morfini (University of Florence), T. Cass (Imperial College), V. Emery (Surrey Business School) and G. Aeppli (Paul Scherrer Institut) for the kind gift of materials and for helpful discussions. The glycoprotein antigens (gp140CN54 and gp140UG37) to llama antibody fragments were provided by the Centre for AIDS Reagents, National Institute for Biological Standards and Control (NIBSC) of the UK Medicines & Healthcare Products Regulatory Agency (MHRA).

Author information

Author notes

  1. Samadhan B. Patil, Manuel Vögtli, Benjamin Webb and Joseph W. Ndieyira: These authors contributed equally to this work

Authors and Affiliations

  1. London Centre for Nanotechnology and Departments of Medicine and Physics, University College London, 17–19 Gordon Street, London, WC1H 0AH, UK

    Samadhan B. Patil, Manuel Vögtli, Benjamin Webb, Rachel A. McKendry & Joseph W. Ndieyira

  2. Department of Materials, Imperial College London, London, SW7 2AZ, UK

    Samadhan B. Patil & Yeong-Ah Soh

  3. Division of Infection & Immunity, University College London, Cruciform Building, Gower Street, London, WC1E 6BT, UK

    Benjamin Webb

  4. UCL Institute for Liver and Digestive Health, Royal Free Hospital, London, NW3 2QG, UK

    Giuseppe Mazza & Massimo Pinzani

  5. Department of Chemistry, Jomo Kenyatta University of Agriculture and Technology, Nairobi, PO Box 62000, Kenya

    Joseph W. Ndieyira

Authors
  1. Samadhan B. Patil
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Manuel Vögtli
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Benjamin Webb
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Giuseppe Mazza
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Massimo Pinzani
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yeong-Ah Soh
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Rachel A. McKendry
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Joseph W. Ndieyira
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

J.W.N. designed the experiments. J.W.N. and M.V. performed the experiments on antibiotics. J.W.N., S.B.P. and B.W. performed the experiments on HIV antigen detections. J.W.N. and S.B.P. formulated the mathematical model to decouple competing surface binding kinetics at Au (top surface of cantilever) and Si (bottom surface of cantilever). J.W.N. performed the experiments on blood clotting proteins and wrote the paper. All authors discussed the results and commented on the manuscript.

Corresponding author

Correspondence to Joseph W. Ndieyira.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary information

Supplementary information (PDF 452 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Patil, S., Vögtli, M., Webb, B. et al. Decoupling competing surface binding kinetics and reconfiguration of receptor footprint for ultrasensitive stress assays. Nature Nanotech 10, 899–907 (2015). https://doi.org/10.1038/nnano.2015.174

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nnano.2015.174

This article is cited by

Search

Advanced 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