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

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.

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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.

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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).

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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.

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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

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